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* Context.-Machine learning applications in the pathology clinical domain are emerging rapidly. As decision support systems continue to mature, laboratories will increasingly need guidance to evaluate their performance in clinical practice. Currently there are no formal guidelines to assist pathology laboratories in verification and/or validation of such systems. These recommendations are being proposed for the evaluation of machine learning systems in the clinical practice of pathology. Objective.-To propose recommendations for performance evaluation of in vitro diagnostic tests on patient samples that incorporate machine learning as part of the pre-analytical, analytical, or post-analytical phases of the laboratory workflow. Topics described include considerations for machine learning model evaluation including risk assessment, predeployment requirements, data sourcing and curation, verification and validation, change control management, human-computer interaction, practitioner training, and competency evaluation. Data Sources.-An expert panel performed a review of the literature, Clinical and Laboratory Standards Institute guidance, and laboratory and government regulatory frameworks. Conclusions.-Review of the literature and existing documents enabled the development of proposed recommendations. This white paper pertains to performance evaluation of machine learning systems intended to be implemented for clinical patient testing. Further studies with real-world clinical data are encouraged to support these proposed recommendations. Performance evaluation of machine learning models is critical to verification and/or validation of in vitro diagnostic tests using machine learning intended for clinical practice. (Arch Pathol Lab Med. 2024;148:e335-e361; doi: 10.5858/arpa.2023-0042-CP), Institution of machine learning (ML) in the pathology clinical domain has gained momentum and is rapidly advancing. As ML-based clinical decision support (CDS) systems continue to mature, laboratories will increasingly [...]

* Context.--Next-generation sequencing (NGS)-based assays are used for diagnosis of diverse inherited disorders. Limited data are available pertaining to interlaboratory analytical performance of these assays. Objective.--To report on the College of American Pathologists (CAP) NGS Germline Program, which is methods based, and explore the evolution in laboratory testing practices. Design.--Results from the NGS Germline Program from 2016-2020 were analyzed for interlaboratory analytical performance. Self-reported laboratory testing practices were also evaluated. Results.--From 2016-2020, a total of 297 laboratories participated in at least 1 program mailing. Of the 289 laboratories that provided information on tests offered, 138 (47.8%) offered only panel testing throughout their enrollment, while 35 (12.1%) offered panels and exome testing, 30 (10.4%) offered only exomes, 9 (3.1%) offered only genomes, and 15 (5.2%) offered panels, exomes, and genomes. The remainder (62 laboratories, 21.4%) changed their test offerings during the 2016-2020 timeframe. Considering each genomic position/interval, the median detection percentage at variant positions across the 2016-2020 mailings ranged from 94.3% to 100%, while at reference positions (no variant detected), the median correct response percentage was 100% across all mailings. When considering performance of individual laboratories, 89.5% (136 of 152) to 98.0% (149 of 152) of laboratories successfully met the detection threshold ([greater than or equal to] 90% of the variants present), while 94.6% (87 of 92) to 100% (163 of 163) of laboratories met the 95% specificity threshold across mailings. Conclusions.--Since the inception of this program, laboratories have consistently performed well. The median sensitivity and specificity of detection of sequence variants included in this program (eg, single nucleotide variants, insertions, and deletions) were 100.0%. doi: 10.5858/arpa.2023-0090-CP), Next-generation sequencing (NGS)-based tests are of higher complexity than most traditional molecular tests owing to the interrogation of many positions simultaneously, as well as requirements for extensive laboratory wet bench [...]

* Context.--The College of American Pathologists (CAP) surveys provide national benchmarks of pathology practice. Objective.--To investigate pancreaticobiliary cytology practice in domestic and international laboratories in 2021. Design.--We analyzed data from the CAP Pancreaticobiliary Cytology Practice Supplemental Questionnaire that was distributed to laboratories participating in the 2021 CAP Nongynecologic Cytopathology Education Program. Results.--Ninety-three percent (567 of 612) of respondent laboratories routinely evaluated pancreaticobiliary cytology specimens. Biliary brushing (85%) was the most common pancreaticobiliary cytology specimen evaluated, followed by pancreatic fine-needle aspiration (79%). The most used sampling methods reported by 235 laboratories were 22-gauge needle for fine-needle aspiration (62%) and SharkCore needle for fine-needle biopsy (27%). Cell block was the most used slide preparation method (76%), followed by liquid-based cytology (59%) for pancreatic cystic lesions. Up to 95% (303 of 320) of laboratories performed rapid on-site evaluation (ROSE) on pancreatic solid lesions, while 56% (180 of 320) performed ROSE for cystic lesions. Thirty-six percent (193 of 530) of laboratories used the Papanicolaou Society of Cytopathology System for Reporting Pancreaticobiliary Cytology in 2021. Among all institution types, significant differences in specimen volume, specimen type, ROSE practice, and case sign-out were identified. Additionally, significant differences in specimen type, slide preparation, and ROSE practice were found. Conclusions.--This is the first survey from the CAP to investigate pancreaticobiliary cytology practice. The findings reveal significant differences among institution types and between domestic and international laboratories. These data provide a baseline for future studies in a variety of practice settings. (Arch Pathol Lab Med. 2024;148:677-685; doi: 10.5858/arpa.2023-0167-CP), Cytology has proven to be a minimally invasive and cost-effective modality for accurately diagnosing pancreaticobiliary lesions, and in some patients, the only modality to diagnose lesions in this anatomic region. [...]

* Context.--Next-generation sequencing-based approaches using RNA have increasingly been used by clinical laboratories for the detection of fusion genes, intragenic rearrangements, and exon-skipping events. Correspondingly, the College of American Pathologists (CAP) has advanced RNA sequencing proficiency testing (PT) to ensure optimal performance of these assays. Objective.--To report on laboratory performance and practices of RNA sequencing for the detection of fusion genes, intragenic rearrangements, and exon-skipping events using CAP PT data from 8 mailings (2018-A through 2021-B). Design.--CAP PT RNA sequencing program results from 153 laboratories across 24 proficiency test specimens, interrogating 22 distinct engineered fusion transcripts, were analyzed for correct identification of the fusion event, associated performance variables, and laboratory practices. Results.--Overall, the 4-year program detection rate (sensitivity) was 95.5% (1486 of 1556 results). False-negative rates were 3.6% (53 of 1463) and 18.3% (17 of 93) for fusion gene and intragenic rearrangement/exon-skipping events, respectively. Only 19 false-positive results were reported among the 8 PT mailings, and most were likely the result of preanalytical or postanalytical errors. There were no practice characteristics (eg, instrumentation, sequencing method) significantly associated with the fusion detection results. Conclusions.--These data reveal a high overall sensitivity and specificity for fusion gene detection by participating laboratories using clinical RNA sequencing. Performance was comparable across all laboratories, regardless of methodology. The fraction of false-negative results for intragenic rearrangement/exon-skipping events was greater than that for the chimeric fusion genes. False-negative results could not be attributed to any specific practice characteristics., Fusion oncogenes represent valuable biomarkers that aid in diagnostic classification and prognostication, as well as serving as therapeutic targets. (1-4) Examples of structural alterations/mutations that may lead to chimeric fusions [...]

* Context.--The Sustainable Predictive Oncology Therapeutics and Diagnostics quality assurance pilot study (SPOT/ Dx pilot) on molecular oncology next-generation sequencing (NGS) reportedly demonstrated performance limitations of NGS laboratory-developed tests, including discrepancies with a US Food and Drug Administration-approved companion diagnostic. The SPOT/Dx pilot methods differ from those used in proficiency testing (PT) programs. Objective.--To reanalyze SPOT/Dx pilot data using PT program methods and compare to PT program data. Design.--The College of American Pathologists (CAP) Molecular Oncology Committee reanalyzed SPOT/Dx pilot data applying PT program methods, adjusting for confounding conditions, and compared them to CAP NGS PT program performance (2019-2022). Results.--Overall detection rates of KRAS and NRAS single-nucleotide variants (SNVs) and multinucleotide variants (MNVs) by SPOT/Dx pilot laboratories were 96.8% (716 of 740) and 81.1% (129 of 159), respectively. In CAP PT programs, the overall detection rates for the same SNVs and MNVs were 97.2% (2671 of 2748) and 91.8% (1853 of 2019), respectively. In 2022, the overall detection rate for 5 KRAS and NRAS MNVs in CAP PT programs was 97.3% (1161 of 1193). Conclusions.--CAP PT program data demonstrate that laboratories consistently have high detection rates for KRAS and NRAS variants. The SPOT/Dx pilot has multiple design and analytic differences with established PT programs. Reanalyzed pilot data that adjust for confounding conditions demonstrate that laboratories proficiently detect SNVs and less successfully detect rare to never-observed MNVs. The SPOT/Dx pilot results are not generalizable to all molecular oncology testing and should not be used to market products or change policy affecting all molecular oncology testing. (Arch Pathol Lab Med. 2024;148:139-148; doi: 10.5858/arpa.2023-0322-CP, Proficiency testing (PT) constitutes a critical component of clinical diagnostics, ensuring laboratories can identify the biomarkers that their validated assays are designed to detect. It is also an essential part [...]

* Context.--Unsatisfactory Papanicolaou (Pap) tests pose a unique set of challenges to the laboratory with regard to their processing, review, reporting, and performance of human papillomavirus (HPV) testing. There are no standardized guidelines for the review process and handling of unsatisfactory Pap tests. Objective.--To assess the current practice patterns regarding various aspects of the unsatisfactory Pap test, from processing to reporting, across laboratories worldwide. Design.--A supplemental questionnaire was mailed to laboratories participating in the 2020 College of American Pathologists (CAP) Gynecologic Cytopathology (PAP Education) Program, requesting data regarding the unsatisfactory Pap test. Results.--Of 1520 participating laboratories, 619 (40.7%) responded, and the responses of 577 laboratories were included for further analysis. Only 64.6% (373 of 577) laboratories used the unsatisfactory Pap test criteria as specified by the 2014 Bethesda System. About three-quarters of the respondents (433 of 576; 75.2%) routinely rescreened unsatisfactory Pap tests. Routine repreparation of such Pap tests was performed by 54.9% (316 of 576) of laboratories, and 52.0% (293 of 563) used glacial acetic acid for repreparing excessively bloody specimens. HPV test results were reported for unsatisfactory Pap tests, always or sometimes, by 62.4% (353 of 566) of respondents. Conclusions.--This CAP survey reveals important information regarding the practice patterns pertaining to several aspects of the unsatisfactory Pap test. It also provides valuable insight into the quality assurance measures that can be implemented for such tests. Future studies can further aid in the standardization of all components of the handling of unsatisfactory Pap tests for overall quality improvement. doi: 10.5858/arpa.2022-0330-CP, The assessment of specimen adequacy or satisfactoriness is a key component of a Papanicolaou (Pap) test interpretation. The criteria for specimen adequacy were initially established under the Bethesda System (TBS) [...]

Context.--Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) assay reagents are available both with and without supplementation with pyridoxal-5'-phosphate (P5P; the active form of vitamin b6), a catalytic cofactor required for their enzymatic reactions. Nonsupplemented assays may miss ALT or AST elevations in patients with vitamin B6 deficiency. Objective.--To assess awareness and adoption of ALT and AST reagents that are supplemented with P5P. Design.--A 4-question survey about ALT and AST reagent supplementation with P5P was included in the College of American Pathologists General Chemistry and Therapeutic Drugs (C program) proficiency testing 2023 B mailing. Results.--Overall, 38% (1651 of 4304) of responding laboratories reported using ALT and/or AST reagent supplemented with P5P. P5P supplementation was more common for nonacademic hospital/medical center laboratories (44%; 713 of 1629) relative to other settings. Of the laboratories that reported not using P5P-supplemented reagents, few (5%; 141 of 2611) cited plans to convert in the future. Despite the availability of P5P-supplemented reagents from several major assay manufacturers, the most common stated barrier for adoption was that the laboratory's reagent manufacturer does not provide P5P-supplemented reagents. Conclusions.--There is a lack of awareness of the existence and benefits of P5P-supplemented ALT and AST reagents. There is a need for ALT and AST assay manufacturers to clarify and standardize the P5P status of ALT and AST reagents. doi: 10.5858/arpa.2024-0097-CP, Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) are liver-associated enzymes commonly used as biomarkers of hepatic injury or disease. Serum or plasma ALT and AST concentrations are typically measured on [...]

Context.--Laboratory testing practices for diagnosis of Clostridioides difficile infection (CDI) have evolved in response to published guidelines, availability of highly sensitive nucleic acid amplification tests (NAATs), perceived problems with the specificity of NAATs, and CDI reporting requirements. Objective.--To assess the current state of laboratory practice for diagnostic CDI testing. Design.--An optional 8-item supplemental questionnaire was distributed in December 2019 to the 1374 laboratories participating in the College of American Pathologists C difficile Detection (CDF) proficiency testing program challenge CDF-C. Results.--Of 1374 CDF-C participants, 1160 (84.4%) responded, predominantly representing laboratories based in the United States (1077 of 1160; 92.8%). The majority reported using a multistep testing algorithm (684 of 1159; 59.0%). Initial testing with a glutamate dehydrogenase and toxin A/B combination test followed by NAAT for discrepant results was the most common testing method (360 of 1146; 31.4%). NAAT alone (299 of 1146; 26.1%) was next, then NAAT followed by an assay that included toxin A/B enzyme immunoassay if NAAT is positive (258 of 1146; 22.5%). Only 5.4% (62 of 1146) reported using toxin A/B immunoassay alone. Most respondents (1093 of 1131; 96.6%) reported rejecting CDI tests on formed stool, but rejection of CDI testing in pediatric patients was uncommon (211 of 1131; 18.7%). Rejection of CDI testing in patients using laxatives was reported more often by US-based respondents (379 of 1054 [36.0%] versus 9 of 77 [11.7%], P < .001). Conclusions.--Multistep algorithms for CDI diagnosis are widely used in line with published recommendations. Most respondents reported rejection of formed stool for CDI testing, but few reported rejection of testing in infants and patients taking laxatives, suggesting these may be areas of opportunity for laboratories to pursue in improving CDI testing practices. doi: 10.5858/arpa.2023-0434-CP, During the last 2 decades, diagnostic Clostridioides difficile infection (CDI) strategies have evolved considerably, but the optimal approach remains unresolved. Cell cytotoxicity neutralization assay is highly sensitive and specific but [...]

* Context.--Substantial variability between different antibody titration methods has been identified since the development and introduction of the uniform procedure in 2008. Objective.--To determine whether more recent methods or techniques decrease interlaboratory and intralaboratory variation measured using proficiency testing. Design.--Proficiency test data for antibody titration between 2014 and 2018 were obtained from the College of American Pathologists. Interlaboratory and intralaboratory variations were compared by analyzing the distribution of titer results by method and phase, comparing the results against the supplier's quality control titer, and by evaluating the distribution of paired titer results when each laboratory received a sample with the same titer twice. Results.--A total of 1337 laboratories participated in the antibody titer proficiency test during the study period. Only 54.1 % (5874 of 10 852) of anti-D and 63.4% (3603 of 5680) of anti-A reported responses were within 1 titer of the supplier's intended result. Review of the agreement between laboratories of the same methodology found that 78.4% (3139 of 4004) for anti-A and 89.0% (9655 of 10 852) of laboratory responses for anti-D fell within 1 titer of the mode response. When provided with 2 consecutive samples of the same titer (anti-D titer: 16), 85% (367 of 434) of laboratories using the uniform procedure and 80% (458 of 576) using the other method reported a titer difference of 1 or less. Conclusions.--Despite advances, interlaboratory and intralaboratory variance for this assay remains high in comparison with the strong reliance on titer results in clinical practice. There needs to be a reevaluation of the role of this test in clinical decision-making. (Arch Pathol Lab Med. 2023;147:1351-1359; doi: 10.5858/arpa.2022-0248-CP), Antibody titration in transfusion medicine is a commonly used clinical assay to detect the strength of reactivity of red cell antibodies present in a patient's plasma. (1) There are a [...]

Context.--Therapy targeted at human epidermal growth factor receptor 2 (HER2; also known as ERBB2) was used initially for breast and gastroesophageal carcinoma and has more recently been adopted for endometrial serous carcinoma (ESC) and colorectal carcinoma (CRC). There is evidence that predictive biomarker testing algorithms for HER2 must be tumor type specific and that an algorithm validated for one tumor type cannot be applied to another. Objective.--To describe current laboratory practices for HER2 assessment in ESC and CRC. Design.--We surveyed laboratories participating in the 2021 College of American Pathologists (CAP) HER2 immunohistochemistry proficiency testing program. Results.--The survey was distributed to 1548 laboratories and returned by 1195, of which 83.5% (998) were in the United States. For ESC, 24.0% (287) of laboratories reported performing in-house testing for HER2 by immunohistochemical staining and/or in situ hybridization; of these, 44.3% (127) performed it reflexively on all cases of ESC. The most common criterion for evaluating HER2 was the American Society of Clinical Oncology/CAP 2018 guideline for breast carcinoma (69.0%; 194 of 281), whereas only 16.0% (45) of laboratories used guidelines specific to ESC. For CRC, 20.2% (239 of 1185) of laboratories performed in-house HER2 testing, and 82.0% of these (196) did the test only at the clinician's request. A plurality (49.4%; 115 of 233) used gastroesophageal cancer guidelines when scoring CRC, 30.0% (70) used the CRC scoring system from the HERACLES trial, and 16.3% (38) used the American Society of Clinical Oncology/CAP 2018 guideline for breast carcinoma. Conclusions.--Laboratories vary in their approach to HER2 testing in ESC and CRC. Most laboratories did not report using tumor type-specific recommendations for HER2 interpretation. The lack of standardization could present a challenge to evidence-based practice when considering targeted therapy for these diseases. doi: 10.5858/arpa.2022-0229-CP, As targeted therapy has become available for an increasing number of tumor types, it has become important to assess predictive biomarkers in the pathology laboratory. For example, the monoclonal antibody [...]

To the Editor.--Syphilis caused by Treponema pallidum subsp pallidum is one of the oldest sexually transmitted diseases, and yet a common one that has made a significant comeback since the [...]

* Context.--Integration of molecular data into glioma classification supports diagnostic, prognostic, and therapeutic decision-making; however, testing practices for these informative biomarkers in clinical laboratories remain unclear. Objective.--To examine the prevalence of molecular testing for clinically relevant biomarkers in adult and pediatric gliomas through review of a College of American Pathologists proficiency testing survey prior to the release of the 2021 World Health Organization Classification of Central Nervous System Tumors. Design.--College of American Pathologists proficiency testing 2020 survey results from 96 laboratories performing molecular testing for diffuse gliomas were used to determine the use of testing for molecular biomarkers in gliomas. Results.--The data provide perspective into the testing practices for diffuse gliomas from a broad group of clinical laboratories in 2020. More than 98% of participating laboratories perform testing for glioma biomarkers recognized as diagnostic for specific subtypes, including IDH. More than 60% of laboratories also use molecular markers to differentiate between astrocytic and oligodendroglial lineage tumors, with some laboratories providing more comprehensive analyses, including prognostic biomarkers, such as CDKN2A/B homozygous deletions. Almost all laboratories test for MGMT promoter methylation to identify patients with an increased likelihood of responding to temozolomide. Conclusions.--These findings highlight the state of molecular testing in 2020 for the diagnosis and classification of diffuse gliomas at large academic medical centers. The findings show that comprehensive molecular testing is not universal across clinical laboratories and highlight the gaps between laboratory practices in 2020 and the recommendations in the 2021 World Health Organization Classification of Central Nervous System Tumors. doi: 10.5858/arpa.2021-0431-CP, Historically, the classification of diffuse gliomas was based in large part on histologic features, including tumor cell morphology, mitotic index, and the presence of necrosis and/or microvascular proliferation. To delineate [...]

Context.--Clinical testing for tumor cell-free DNA (cfDNA) has evolved rapidly, but no practice guidelines exist. Objective.--To summarize cfDNA laboratory practices based on self-reporting and assess preanalytical, analytical, and postanalytical trends that may influence the quality, accuracy, and consistency of cfDNA testing. Design.--Data were derived from the College of American Pathologists cfDNA proficiency testing program submitted by 101 participating laboratories from 2018 to 2019. Results.--Most laboratories performing clinical circulating tumor DNA testing are commercial/nonhospital (71.2%; 72 of 101) and international (77.2%; 78 of 101) laboratories. Commercial laboratories had higher monthly test volumes than hospital-based laboratories (median, 36 versus 7-8) and tended to have larger gene panels (median, 50 versus 11 genes) when panel-based testing was offered. The main clinical indications include therapy selection and treatment/disease monitoring. Plasma is the most commonly accepted specimen, which is predominantly collected in cell-stabilizing tubes. Equal proportions of laboratories use next-generation sequencing (NGS) and non-NGS methods to assess key genes, including EGFR, BRAF, KRAS, NRAS, and IDH1. Most laboratories reported a lower limit of detection (LLOD) of 0.5%, variant allele frequency or less, which did not differ by method, NGS or non-NGS, except for EGFR. Sixty-five percent (17 of 26) of laboratories using the US Food and Drug Administration (FDA)-approved non-NGS EGFR assay report analytical sensitivities higher than 0.5%, as compared to 15% (16 of 104) of laboratories using an alternative NGS or non-NGS method. There is also a wider range in LLODs obtained for the FDA-approved EGFR assay than nonapproved assays. Conclusions.--These results highlight emerging practice trends and serve as a foundation to initiate future practice recommendations. doi: 10.5858/arpa.2021-0585-CP, Cell-free DNA (cfDNA) was first reported in whole blood in 1948, which consists of short fragments of circulating DNA that are approximately 150 to 200 base pairs in length. (1,2) [...]

* Context.--Most laboratories currently use patient tissues for validating immunohistochemical stains. Objective.--To explore advantages of using cell lines with known antigenicity as a validation method. Design.--Five American Type Culture Collection (ATCC) cell lines with known negative, low positive, and moderate to strong estrogen receptor (ER) expression as well as negative, equivocal, and positive human epidermal growth factor receptor 2 (HER2) expression were cultured and made into cell blocks. One block from each cell line was fixed in formalin and another in ethanol before cell block preparation. Two sets of paired unstained slides from each block were sent to 10 different laboratories for HER2 and ER staining to be stained on runs from different days according to each laboratory's defined protocol. Results.--The 10 study participants evaluated 40 slides in a blinded fashion. For ER expression, all 80 interpretations for the ER strong and moderate positive cell lines had the target ER-positive result, and 74 of 80 ER-negative cell lines (92.5%) had agreement with the intended negative result. The ER low positive cell line showed varied but positive expression among all observers. The HER2 (3+)positive cell lines yielded a target interpretation of 3+ in 65 of 80 interpretations (81.2%). For the HER2-negative cell line 69 of 78 interpretations (88.5%) were consistent with the target response (0 or 1+). No significant variation was observed between the ethanol- and non-ethanol-exposed cell lines, or between runs by the same laboratory. Variation from target results clustered within laboratories. Conclusions.--This study indicates that variability between laboratories can be identified by using cell lines for quantitative or semiquantitative immunohistochemistry when using cultured cell lines of known antigenicity. These cell lines could potentially play a role in aiding anatomic pathology laboratories in validating immunohistochemistry tests for formalin- and ethanol-fixed tissues. doi: 10.5858/arpa.2021-0152-CP, Reliable immunohistochemical stain results are often a needed aid in the diagnostic assessment of anatomic pathology cases. Furthermore, as the sophistication of tumor subtyping has continued to evolve, most IHC [...]

* Context.--In 2016, the College of American Pathologists (CAP) launched the first next-generation sequencing (NGS) in silico bioinformatics proficiency testing survey to evaluate the performance of clinical laboratory bioinformatics pipelines for the detection of oncology-associated variants at varying allele fractions. This survey focused on 2 commonly used oncology panels, the Illumina TruSeq Amplicon Cancer Panel and the Thermo Fisher Ion AmpliSeq Cancer Hotspot v2 Panel. Objective.--To review the analytical performance of laboratories participating in the CAP NGS bioinformatics (NGSB) surveys, comprising NGSB1 for Illumina users and NGSB2 for Thermo Fisher Ion Torrent users, between 2016 and 2019. Design.--Responses from 78 laboratories were analyzed for accuracy and associated performance characteristics. Results.--The analytical sensitivity was 90.0% (1901 of 2112) for laboratories using the Illumina platform and 94.8% (2153 of 2272) for Thermo Fisher Ion Torrent users. Variant type and variant allele fraction were significantly associated with performance. False-negative results were seen mostly for multi-nucleotide variants and variants engineered at variant allele fractions of less than 25%. Analytical specificity for all participating laboratories was 99.8% (9303 of 9320). There was no statistically significant association between deletion-insertion length and detection rate. Conclusions.--These results demonstrated high analytical sensitivity and specificity, supporting the feasibility and utility of using in silico mutagenized NGS data sets as a supplemental challenge to CAP surveys for oncology-associated variants based on physical samples. This program demonstrates the opportunity and challenges that can guide future surveys inclusive of customized in silico programs. doi: 10.5858/arpa.2021-0384-CP, Next-generation sequencing (NGS) technologies have enabled comprehensive assessment of tumor molecular profiles and revolutionized the clinical testing landscape in oncology. In addition to allowing the assessment of nucleic acid in [...]

* Context.--Modern RHD genotyping can be used to determine when patients with serologic weak D phenotypes have RHD gene variants at risk for anti-D alloimmunization. However, serologic testing, RhD interpretations, and laboratory management of these patients are quite variable. Objective.--To obtain interlaboratory comparisons of serologic testing, RhD interpretations, Rh immune globulin (RhIG) management, fetomaternal hemorrhage testing, and RHD genotyping for weak D-reactive specimens. Design.--We devised an educational exercise in which 81 transfusion services supporting obstetrics performed tube-method RhD typing on 2 unknown red blood cell challenge specimens identified as (1) maternal and (2) newborn. Both specimens were from the same weak D-reactive donor. The exercise revealed how participants responded to these different clinical situations. Results.--Of reporting laboratories, 14% (11 of 80) obtained discrepant immediate-spin reactions on the 2 specimens. Nine different reporting terms were used to interpret weak D-reactive maternal RhD types to obstetricians. In laboratories obtaining negative maternal immediate-spin reactions, 28% (16 of 57) performed unwarranted antiglobulin testing, sometimes leading to recommendations against giving RhIG. To screen for excess fetomaternal hemorrhage after a weak D-reactive newborn, 47% (34 of 73) of reporting laboratories would have employed a contraindicated fetal rosette test, risking false-negative results and inadequate RhIG coverage. Sixty percent (44 of 73) of laboratories would obtain RHD genotyping in some or all cases. Conclusions.--For obstetric and neonatal patients with serologic weak D phenotypes, we found several critical problems in transfusion service laboratory practices. We provide recommendations for appropriate testing, consistent immunohematologic terminology, and RHD genotype-guided management of Rh immune globulin therapy and RBC transfusions., In the United States and Canada, 0.06% to 2.2% of patients have red blood cells (RBCs) that type weakly reactive for RhD, termed 'serologic weak D phenotype.' (1-6) In past [...]

* Context.--The College of American Pathologists (CAP) updated the Laboratory Accreditation Program Cytopathology Checklist to assist laboratories in meeting and exceeding the Clinical Laboratory Improvement Amendments standards for gynecologic cytologic-histologic correlation (CHC). Objective.--To survey the current CHC practices. Design.--Data were analyzed from a survey developed by the committee and distributed to participants in the CAP Gynecologic Cytopathology PAP Education Program mailing. Results.--Worldwide, CHC practice is nearly universally adopted, with an overall rate of 87.0% (568 of 653). CHC material was highly accessible. CHC was commonly performed real time/concurrently at the time the corresponding surgical pathology was reviewed. Investigation of CHC discordances varied with North American laboratories usually having a single pathologist review all discrepant histology and cytology slides to determine the reason for discordance, while international laboratories have a second pathologist review histology slides to determine the reason for discordance. The cause of CHC discordance was primarily sampling issues. The more common statistical metrics for CHC monitoring were the total percentage of cases that correlated with subsequent biopsies, screening error rate by cytotechnologist, and interpretative error rate by cytotechnologist. Conclusions.--Many laboratories have adopted and implemented the CHC guidelines with identifiable differences in practices between North American and international laboratories. We identify the commonalities and differences between North American and international institutional practices including where CHC is performed, how CHC cases are identified and their accessibility, when CHC is performed, who investigates discordances, what discordances are identified, and how the findings affect quality improvement., The Clinical Laboratory Improvement Amendments (CLIA) were passed in 1988 and laboratories were asked to comply with certain gynecologic testing standards, requiring 'laboratory comparison of clinical information, when available, with [...]

* Context.--Neoplastic cellularity assessment has become an essential component of molecular oncology testing; however, there are currently no best practice recommendations or guidelines for this potentially variable step in the testing process. Objective.--To describe the domestic and international practices of neoplastic cellularity assessment and to determine how variations in laboratory practices affect neoplastic cellularity assessment accuracy. Design.--Data were derived from 57 US and international laboratories that participated in the 2019 College of American Pathologists Neoplastic Cellularity Proficiency Testing Survey (NEO-B 2019). NEO-B 2019 included 29 laboratory practice questions and 5 images exhibiting challenging histologic features. Participants assessed the neoplastic cellularity of hematoxylin-eosin-stained digital images, and results were compared to a criterion standard derived from a manual cell count. Results.--The survey responses showed variations in the laboratory practices for the assessment of neoplastic cellularity, including the definition of neoplastic cellularity, assessment methodology, counting practices, and quality assurance practices. In some instances, variation in laboratory practice affected neoplastic cellularity assessment performance. Conclusions.--The results highlight the need for a consensus definition and improved standardization of the assessment of neoplastic cellularity. We put forth an initial set of best practice recommendations to begin the process of standardizing neoplastic cellularity assessment. (Arch Pathol Lab Med. 2022;146:1062-1071; doi: 10.5858/arpa.2021-0166-CP), In the era of precision medicine, molecular oncology testing has become critical to identifying clinically relevant mutations in a variety of cancer genes that are used in diagnosis, prognosis, and/or [...]

Context.--The 2017 Association for Molecular Pathology/American Society of Clinical Oncology/College of American Pathologists (CAP) tier classification guideline provides a framework to standardize interpretation and reporting of somatic variants. Objective.--To evaluate the adoption and performance of the 2017 guideline among laboratories performing somatic next-generation sequencing (NGS). Design.--A survey was distributed to laboratories participating in NGS CAP proficiency testing for solid tumors (NGSST) and hematologic malignancies (NGSHM). Results.--Worldwide, 64.4% (152 of 236) of NGSST and 66.4% (87 of 131) of NGSHM participants used tier classification systems, of which the 2017 guideline was used by 84.9% (129 of 152) of NGSST and 73.6% (64 of 87) of NGSHM participants. The 2017 guideline was modified by 24.4% (30 of 123) of NGSST and 21.7% (13 of 60) of NGSHM laboratories. Laboratories implementing the 2017 guideline were satisfied or very satisfied (74.2% [89 of 120] NGSST and 69.5% [41 of 59] NGSHM), and the impression of tier classification reproducibility was high (mean of 3.9 [NGSST] and 3.6 [NGSHM] on a 5-point scale). Of nonusers, 35.2% (38 of 108) of NGSST and 39.4% (26 of 66) of NGSHM laboratories were planning implementation. For future guideline revisions, respondents favored including variants to monitor disease (63.9% [78 of 122] NGSST, 80.0% [48 of 60] NGSHM) and germline variants (55.3% [63 of 114] NGSST, 75.0% [45 of 60] NGSHM). Additional subtiers were not favored by academic laboratories compared to nonacademic laboratories (P Conclusions.--The 2017 guideline has been implemented by more than 50.0% of CAP laboratories. While most laboratories using the 2017 guideline report satisfaction, thoughtful guideline modifications may further enhance the quality, reproducibility, and clinical utility of the 2017 guideline for tiered somatic variant classification., The increasing availability of next-generation sequencing (NGS) of cancers has led to an exponential increase in the amount and complexity of molecular data generated in clinical settings. (1) To provide [...]

* Context.--The yield of the prospective rescreening process for 'negative for intraepithelial lesion or malignancy' (NILM) Papanicolaou (Pap) tests is higher with the inclusion of a greater proportion of high-risk cases. One of the suggested criteria for classifying a Pap test finding as high risk is recent or concurrent high-risk human papillomavirus (HPV) positivity. Objective.--To evaluate how the results of HPV testing have been incorporated in the prospective rescreening of NILM Pap tests across a wide range of laboratories. Design.--A questionnaire survey was sent to laboratories participating in the 2019 College of American Pathologists (CAP) Gynecologic Cytology (PAP Education) Program. Results.--Of the 1507 participating laboratories, 667 (44%) responded to the survey. Most laboratories (59.4%; 396 of 667) had not incorporated HPV test/genotyping results to select NILM Pap tests for rescreening. Amongst the remaining laboratories, for NILM HPV-positive Pap test results, 112 (16.8%) had a policy to rescreen by a cytotechnologist only, 51 (7.6%) by a pathologist only, and 86 (12.9%) by both. Of 264 laboratories, 181 (68.6%) reported the cytology upon availability of the HPV test result and completion of the secondary review. Of 661 laboratories, 145 (21.9%) included consensus-type recommendations in the cytology report for such Pap tests. Conclusions.--This CAP survey provides significant information regarding the current trends in the use of HPV test results in prospective rescreening of NILM Pap tests. Future studies on quality improvement can further assist in the standardization of this process across different laboratories., The practice of gynecologic cytology is a highly regulated field. The Clinical Laboratory Improvement Amendments of 1988 (CLIA) set several mandates for gynecologic cytology. (1) These included the prospective rescreening [...]

* Context.--The College of American Pathologists surveys provide national benchmarks of pathology practice for laboratories. Objective.--To investigate breast fine-needle aspiration (FNA) biopsy practice in domestic and international laboratories in 2019. Design.--We analyzed data from the College of American Pathologists Breast FNA Practice Supplemental Questionnaire that was distributed to laboratories participating in the 2019 College of American Pathologists Non-Gynecologic Cytopathology Education Program. Results.--Sixty-one percent (499 of 816) of respondent laboratories routinely evaluated breast FNAs. Cystic lesions were the most common indication, and radiologists primarily performed FNAs in most settings. Forty-five percent (220 of 491) of laboratories performed ancillary studies on breast FNA samples, but 33.8% (70 of 207) did not report fixation time for breast biomarker studies. Only 54.5% (271 of 497) of laboratories had a standardized reporting system and only 16.8% (82 of 488) were aware of the International Academy of Cytology Yokohama Breast FNA Biopsy Cytology Reporting System. There were significant differences among different types of institutions in several aspects of breast FNA practice, including frequency of concurrent FNA and core needle biopsy for the same lesion, primary personnel who performed the FNA, etc. Significant differences existed between domestic and international laboratories in slide preparation, ancillary studies, fixation time reporting, standardized/descriptive diagnosis, and International Academy of Cytology Yokohama Reporting System awareness. Conclusions.--This is the first survey from the College of American Pathologists Cytopathology Committee to investigate breast FNA practices. The data reveal significant differences in breast FNA practice among different types of institutions and between domestic and international laboratories, and provide a baseline for future breast FNA studies in a variety of practice settings. (Arch Pathol Lab Med. 2021;145:825-833; doi: 10.5858/arpa.2020-0408-CP), Breast fine-needle aspiration (FNA) biopsy is performed with variable frequency among different countries and institutions. (1-9) In 1996, a National Cancer Institute-sponsored conference with a group of breast practitioners put [...]

ContextThe College of American Pathologists periodically surveys laboratories to determine changes in cytopathology practices. We report the results of a 2011 gynecologic cytology survey.ObjectiveTo provide a cross-sectional survey of gynecologic cytology practices in 2010.DesignIn 2011, a survey was sent to 1604 laboratories participating in the College of American Pathologists gynecologic cytology interlaboratory comparison education program and proficiency testing programs requesting data from 2010 on the following topics: terminology/reporting, cytotechnologist workload, quality assurance, reagents, and ancillary testing.ResultsSix hundred and twenty-five laboratories (39%) replied to the survey. The nonstandard use of "low-grade squamous intraepithelial lesion cannot exclude high-grade squamous intraepithelial lesion" is used by most laboratories to report the presence of low-grade squamous intraepithelial lesion with possibility of high-grade squamous intraepithelial lesion. Most laboratories also report the presence or absence of cells from the transformation zone. Most respondents do not limit cytotechnologist screening workload during the work shift. Only about one-third of laboratories (188 of 582; 32%) use image-assisted screening devices. Rapid prescreening as a quality assurance measure is used by only 3.5% (21 of 594) of the laboratories. When used for screening, most laboratories use the imager for retrospective review of slides to detect human locator and interpretive errors. Most laboratories receive both liquid-based cytology samples (mainly ThinPrep, Hologic, Marlborough, Massachusetts) and conventional Papanicolaou tests. Expiration dates of liquid-based cytology test vials are not usually recorded.ConclusionsThe field of gynecologic cytology is evolving rapidly. These survey results offer a snapshot of national gynecologic cytology practices in 2010.

* Context.--Repair is a challenging diagnosis and a significant source of false-positive (FP) interpretations in cervical cytology. No large-scale study of performance of repair in the liquid-based era has been performed. Objective.--To evaluate the performance of repair in the College of American Pathologists Pap Education and Proficiency Testing (PT) programs. Design.--The FP rate for slides classified as repair was evaluated by preparation type, participant type (cytotechnologist, pathologist, or laboratory), and program. The specific misdiagnosis category and individual slide performance were also evaluated. The rate of misclassification of slides as repair by participants for other diagnostic categories in the Pap Education program was assessed. Results.--The overall FP rate was 1700 of 12 715 (13.4%). There was no significant difference by program or preparation type. Within the Education program there was no difference by participant type, but pathologists' FP rate in the PT program (47 of 514, 9.1%) was significantly better than cytotechnologists in the PT program (51 of 380, 13.4%) and pathologists in the Education program (690 of 4900, 14.1%). High-grade squamous intraepithelial lesions/cancers ([HSIL.sup.+]) accounted for 1380 of 1602 FP interpretations (86%) in Education, but 43 of 98 (43.9%) in PT. Most slides had a low rate of misclassification, but a small number were poor performers. False-negative diagnosis of [HSIL.sup.+] as repair was less common, ranging from 0.7% to 1.8%. Conclusions.--Despite initial indications that liquid-based cytology might reduce the rate of misclassification of repair, FP interpretations remain common and are no different by preparation type. Misclassification is most commonly as HSIL or carcinoma, potentially resulting in significant patient harm. (Arch Pathol Lab Med. 2020;144:846-852; doi: 10.5858/arpa.2019-0298-CP), Repair, or reparative changes, refers to a specific pattern of changes in cervical cells attributed to injury from an external stimulus. (1-4) Although repair shows some morphologic overlap with dysplastic [...]

* Context.--Consolidation of clinical microbiology laboratory services has resulted in extended transit time for blood cultures from service points distant from the laboratory. Sepsis is critical; delays in identification of etiologic agents of diseases could adversely impact patient care. Objective.--To examine the effect of total preanalytic time and blood culture volume on the instrument time-to-detection for bacterial pathogens in blood cultures. A secondary objective was to obtain relevant blood culture information by questionnaire. Design.--Participants in this Q-Probes study recorded date, time, and volume information for the first 50 positive blood cultures collected during the 12-week study period. Additional information regarding blood culture collection practices was obtained through questionnaire. Results.--Prolonged overall time-to-detection was secondary to prolonged preanalytic time, particularly prolonged transit time, rather than slower organism growth once bottles were placed on the instrument. Among 1578 blood cultures, the overall time from collection to positive result was significantly less for blood cultures collected onsite than for off-site locations. Most institutions lack sufficient training programs and do not monitor preanalytic time metrics associated with blood cultures. Four hundred fifty-six of the 1580 blood cultures with blood volume adequacy reported (28.9%) were inadequately filled. Conclusions.--Overall process time (specimen collection to positive blood culture detection) is predicted to be higher for blood cultures collected off-site. Transit time is a variable that can be reduced to decrease overall time to detection. Thus, improved training and closer attention to preanalytic metrics associated with blood cultures could decrease hospital stays and mortality rates. (Arch Pathol Lab Med. 2020;144:564-571; doi: 10.5858/arpa.2019-0258-CP), The incidence of sepsis and septic shock has increased in the United States, with more than 1.5 million people developing sepsis each year in the United States and about 250 [...]

Context.--Detection of high-risk human papillomavirus (HR-HPV) in squamous cell carcinoma is important for classification and prognostication. In situ hybridization (ISH) is a commonly used HR-HPV-specific test that targets viral RNA or DNA. The College of American Pathologists (CAP) provides proficiency testing for laboratories performing HR-HPV iSh. Objective.--To compare the analytical performance of RNA- and DNA-based ISH methods on CAP HR-HPV proficiency tests. Design.--Data from the 2016-2018 CAP HPV ISH proficiency testing surveys were reviewed. These surveys consist of well-characterized samples with known status for HR-HPV, including 1 to 2 copies, 50 to 100 copies, 300 to 500 copies, and no copies of HR-HPV per cell. Results.--Ninety-five participants submitted 1268 survey results from 20 cores. Overall, RNA ISH had a significantly higher percentage of correct responses than DNA ISH: 97.4% (450 of 462) versus 80.6% (650 of 806) (P < .001). This disparity appears to be the consequence of a superior sensitivity of rNa ISH compared to DNA ISH for samples with 1 to 2 and with 50 to 100 copies of HRHPV per cell: 95.2% (120 of 126) versus 53.8% (129 of 240), P < .001, respectively, and 100% (89 of 89) versus 76.3% (119 of 156), P < .001, respectively. Conclusions.--An assessment of CAP HR-HPV proficiency test performance indicates that RNA ISH shows significantly higher accuracy than DNA ISH owing to higher analytical sensitivity of RNA ISH in tumors with low (1-2 copies per cell) to intermediate (50-100 copies per cell) HR-HPV viral copy numbers. These data support the use of RNA over DNA ISH in clinical laboratories that perform HR-HPV testing as part of their testing algorithms. (Arch Pathol Lab Med. 2020;144:344-349; doi: 10.5858/arpa.2019-0093-CP), High-risk human papillomavirus (HR-HPV) causes HPV-positive squamous cell carcinomas (SCCs) at multiple anatomic sites including the oropharynx, anus, and urogenital tracts. The detection of HR-HPV in tissue samples containing SCC [...]

* Context.--The Paris System for Reporting Urinary Cytology has been disseminated since its inception in 2013; however, the daily practice patterns of urinary tract cytopathology are not well known. Objective.--To assess urinary tract cytopathology practice patterns across a variety of pathology laboratories to aid in the implementation and future update of the Paris System for Reporting Urinary Cytology. Design.--A questionnaire was designed to gather information about urinary tract cytopathology practices and mailed in July 2014 to 2116 laboratories participating in the College of American Pathologists interlaboratory comparison program. The participating laboratories' answers were summarized. Results.--Of the 879 of 2116 laboratories (41%) that participated, 745 (84.8%) reported processing urinary tract specimens in house. The laboratories reported processing various specimen types: voided urine, 735 of 738 (99.6%); bladder washing/barbotage, 639 of 738 (86.6%); and catheterized urine specimens, 653 of 738 (88.5%). Some laboratories used multiple preparation methods, but the most commonly used preparation techniques for urinary tract specimens were ThinPrep (57.4%) and Cytospin (45.5%). Eighty-eight of 197 laboratories (44.7%) reported preparing a cell block, but with a low frequency. Adequacy criteria were used by 295 of 707 laboratories (41.7%) for voided urine, and 244 of 707 (34.5%) assessed adequacy for bladder washing/ barbotage. More than 95% of the laboratories reported the use of general categories: negative, atypical, suspicious, and positive. Polyomavirus was classified as negative in 408 of 642 laboratories (63.6%) and atypical in 189 of 642 (29.4%). One hundred twenty-eight of 708 laboratories (18.1%) performed ancillary testing, and of these, 102 of 122 (83.6%) reported performing UroVysion. Conclusions.--Most laboratories use the ThinPrep method followed by the Cytospin technique; therefore, the criteria published in The Paris System for Reporting Urinary Cytology, based mostly on ThinPrep and SurePath, should be validated for Cytospin, and relevant information should be included in the revised edition of The Paris System for Reporting Urinary Cytology. (Arch Pathol Lab Med. 2020;144:172-176; doi: 10.5858/arpa.2019-0045-CP), Urine cytology comprises a variable, albeit significant, percentage of daily nongynecologic case volume in many cytopathology practices and continues to be one of the more difficult specimens that pathologists encounter [...]

* Context.--To date, the College of American Pathologists (CAP) has developed 17 laboratory practice guidelines (LPGs) including updates. In 2013, the CAP was awarded a 5-year cooperative agreement grant from the United States Centers for Disease Control and Prevention to increase the effectiveness of LPGs. Objective.--To assess the awareness and adoption of 2 CAP LPGs: immunohistochemical (IHC) assay validation and initial workup of acute leukemia. Design.--Baseline surveys for each LPG were conducted in 2010 and 2015, respectively. To measure the adoption of guideline recommendations and inform future updates, a follow-up study consisting of surveys, telephone interviews, and focus group sessions was conducted in laboratories that indicated they perform IHC testing. A follow-up study for the acute leukemia LPG is planned. Results.--For the IHC Validation LPG, a total of 1624 survey responses, 40 telephone interviews, and discussions with 5 focus group participants were analyzed. The response rate for the aforementioned 3 modalities was 46%, 13%, and 3%, respectively. All modalities indicated most respondents were aware of the LPG and had adopted most or all of its recommendations. Respondents expressed needs for continued communication, increased specificity, and more prescriptive recommendations when the guideline is updated. Conclusions.--While data-driven development of evidence-based LPGs requires significant resources, active data collection to identify gaps and assess adoption contributes to improved laboratory testing practices in support of patient care. The CAP identified sustainable modalities to track metrics and developed multiple tools that should improve guideline development, adoption, and implementation. Of these modalities, written or electronic surveys were the most logistically feasible and had the highest response rate. (Arch Pathol Lab Med. 2020;144:83-89; doi: 10.5858/arpa.2018-0276-CP), The United States Clinical Laboratory Improvement Amendments of 1988 (CLIA '88) established minimum requirements for assuring the quality of laboratory testing in US clinical laboratories. (1) To ensure accurate diagnosis [...]

ContextIn cytology proficiency testing (PT), participants fail for incorrectly interpreting a high-grade squamous intraepithelial lesion or cancer (HSIL+) Papanicolaou test result as negative. This penalty may lead to a false-positive interpretation of negative slides as HSIL+ to avoid failure.ObjectiveTo investigate factors related to false-positive responses in a PT versus an educational environment.DesignWe analyzed 420,079 responses from 9414 validated negative reference slides in the College of American Pathologists Interlaboratory Comparison Program in Gynecologic Cytopathology (PAP Education) and compared them with responses from the Gynecologic Cytology Proficiency Testing Program for the percentage of false-positive (HSIL+) interpretations in each of 7 negative subcategories. We evaluated the influence of preparation type (ThinPrep, SurePath, and conventional Papanicolaou test), participant type (pathologist or cytotechnologist), and program time interval (preproficiency test or PT) on a false-positive response.ResultsReference diagnosis and participant type, but not preparation type, were statistically correlated to false-positive responses. The interaction between program time interval and participant type was also significant. Pathologists had higher rates of false-positive results on preproficiency test (1.2% [800 of 68,690]) than they did on PT (0.8% [993 of 129,857]). Cytotechnologists had no differences between program time intervals (preproficiency, 0.9% [515 of 63,281] versus PT, 1.0 [1231 of 121,621]; P = .91). Negative subcategories frequently mistaken for HSIL+ were reparative changes (4.7% [427 of 9069]), atrophic vaginitis (1.8% [18 of 987]), and negative for intraepithelial lesion or malignancy (1.2% [2143 of 178,651]), but during PT, false-positive rates were significantly increased only for the negative for intraepithelial lesion or malignancy and herpes simplex virus (P < .001).ConclusionsPathologists had lower false-positive rates in the Gynecologic Cytology Proficiency Testing Program than they did in PAP Education, but participants were more likely to report a false-positive response (HSIL+) for negative for intraepithelial lesion or malignancy and herpes simplex virus in the Gynecologic Cytology Proficiency Test Program.

Context.--The Kleihauer-Betke (KB) test is validated for estimating the dose of Rh immune globulin needed for Rh-negative pregnant females. However, some clinicians are also ordering the test for Rh-positive women. The degree to which this practice occurs is unknown. Objective.--To evaluate the number of laboratories that perform the KB test on Rh-positive pregnant women, and to establish current ordering practices for this indication. Design.--We added 9 supplemental questions regarding KB test use for fetomaternal hemorrhage to the 2016 College of American Pathologists proficiency test survey. We also reviewed the available literature regarding the diagnostic utility of the KB test for Rh-positive women. Results.--A total of 1578 surveys were evaluated and revealed that 52% (824) of respondents perform these tests for Rh-positive women, and more than 50% (440 of 819; 53.7%) of these laboratories report that the results for Rh-positive women are treated as important or very important. Conclusions.--The KB test is commonly used for Rh-positive women, and the information obtained from the test is considered as urgent and important. However, the available literature in support of this practice is still nonconclusive. doi: 10.5858/arpa.2018-0523-CP, In the United States, nearly all (96.0%) quantitative assays for fetal red blood cells in maternal circulation are performed using several commercially marketed kits for an acid-elution assay, also known [...]

* Context.--The College of American Pathologists periodically surveys laboratories to determine changes in cytopathology practices. We report the results of a 2016 survey concerning thyroid fine-needle aspiration (FNA). Objective.--To provide a cross-sectional survey of thyroid cytology practices in 2016. Design.--In 2016, a survey was sent to 2013 laboratories participating in the College of American Pathologists Non-Gynecologic Cytology Education Program (NGC-A) requesting data from 2015-2016 on several topics relating to thyroid FNA. Results.--A total of 878 laboratories (43.6% of 2013) replied to the survey. Radiologists performed the most thyroid FNA procedures in most laboratories (70%; 529 of 756), followed by endocrinologists (18.7%; 141 of 756), and most of these were performed under ultrasound guidance (92.1%; 699 of 759). A total of 32.6% of respondents (251 of 769) provided feedback on unsatisfactory rates for nonpathology providers who performed FNA. Intraprocedural adequacy assessment was primarily performed by attending pathologists (77.4%; 490 of 633) or cytotechnologists (28.4%; 180 of 633). Most laboratories used the Bethesda System for Reporting Thyroid Cytopathology (89.8%; 701 of 781) and performed molecular testing based on clinician request (68.1%; 184 of 270) rather than FNA diagnosis. Correlation of thyroid excisions with prior cytology results most often occurred retrospectively (38.4%; 283 of 737) and was used for pathologist interpretive quality assurance purposes. Conclusions.--These survey results offer a snapshot of national thyroid FNA cytology practices in 2016 and indicate that standardized cytology terminology is commonly used; pathologists perform most immediate adequacy assessments for thyroid FNA; laboratories use correlation statistics to evaluate pathologists' performance; and molecular tests are increasingly requested for indeterminate interpretations, but reflex molecular testing is rare. (Arch Pathol Lab Med. 2019;143:1364-1372; doi: 10.5858/arpa.2018-0429-CP), As in decades past, fine-needle aspiration (FNA) remains the primary means to assess the nature of thyroid nodules. Thyroid FNA has been a resounding success, resulting in reduced thyroidectomy rates, [...]

* Context.--Cervical cancer screening laboratory practices may evolve with new terminology and technologies. Objective.--To investigate changes in cervical cytopathology practice resulting from the 2014 Bethesda System updates and screening technologies. Design.--Questionnaires accompanied 2016 and 2017 mailings of the College of American Pathologists PAP Education program. Results.--In 2016, most laboratories surveyed had adopted or were planning to adopt 2014 Bethesda System updates, and the majority (53%; 365 of 689) used an age cutoff of 45 for reporting benign-appearing endometrial cells. However, 51.3% (354 of 690) of laboratories used the term low-grade squamous intraepithelial lesion, cannot exclude high-grade squamous intraepithelial lesion, for cases with indeterminate features, and 44.9% (298 of 664) of laboratories used a 5000-cell cutoff for minimum squamous cellularity for posthysterectomy and posttherapy specimens. Reporting rates for cervical cytology metrics changed very little from 2013 to 2016, and the median ratio of atypical squamous cells to squamous intraepithelial lesion cases was 1.9 for ThinPrep and 1.8 for SurePath preparations. Most laboratories (59.4%; 389 of 655) did not offer stand-alone primary human papillomavirus (HPV) testing in 2017, and primary HPV testing accounted for a low proportion of HPV testing volumes. The Roche Cobas method was the most common platform for HPV primary screening. Conclusions.--These questionnaire surveys provide data about the current status of cervical cytology screening, including changes related to the 2014 Bethesda System updates and the adoption of HPV primary screening techniques. (Arch Pathol Lab Med. 2019;143:1196-1202; doi: 10.5858/arpa.2018-0603-CP), The College of American Pathologists (CAP) has conducted regular questionnaire surveys that assess cytopathology laboratory practices of laboratories participating in the CAP Gynecologic Cytopathology PAP Education program. (1-7) Early CAP [...]

* Context.--Obtaining diagnostic concordance for squamous intraepithelial lesions in cytology can be challenging.Objective.--To determine diagnostic concordance for biopsy-proven low-grade squamous intraepithelial lesion (LSIL) and high-grade squamous intraepithelial lesion (HSIL) Papanicolaou test slides in the College of American Pathologists PAP Education program.Design.--We analyzed 121 059 responses from 4251 LSIL and HSIL slides for the interval 2004 to 2013 using a nonlinear mixed-model fit for reference diagnosis, preparation type, and participant type. We evaluated interactions between the reference diagnosis and the other 2 factors in addition to a repeated-measures component to adjust for slide-specific performance.Results.--There was a statistically significant difference between misclassification of LSIL (2.4%; 1384 of 57 664) and HSIL (4.4%; 2762 of 63 395). There was no performance difference between pathologists and cytotechnologists for LSIL, but cytotechnologists had a significantly higher HSIL misclassification rate than pathologists (5.5%; 1437 of 27 534 versus 4.0%; 1032 of 25 630; P = .01), and both were more likely to misrepresent HSIL as LSIL (P < .001) than the reverse. ThinPrep LSIL slides were more likely to be misclassified as HSIL (2.4%; 920 of 38 582) than SurePath LSIL slides (1.5%; 198 of 13 196), but conventional slides were the most likely to be misclassified in both categories (4.5%; 266 of 5886 for LSIL, and 6.5%; 573 of 8825 for HSIL).Conclusions.--More participants undercalled HSIL as LSIL (false-negative) than overcalled LSIL as HSIL (false-positive) in the PAP Education program, with conventional slides more likely to be misclassified than ThinPrep or SurePath slides. Pathologists and cytotechnologists classify LSIL equally well, but cytotechnologists are significantly more likely to undercall HSIL as LSIL than are pathologists.doi: 10.5858/arpa.2018-0003-CP), The College of American Pathologists (CAP) Cytopathology Committee maintains and administers the PAP Education (PAP Ed) program, which provides 2 mailings of 5 glass slide Papanicolaou (Pap) tests annually to [...]

ContextAnal cytology is being used more frequently for anal cancer screening, yet many cytologists are unfamiliar with it.ObjectiveTo describe the performance of anal cytology in the College of American Pathologists' Interlaboratory Comparison Program in Non-Gynecologic Cytology (CAP NGC) educational slide program during a 6-year time span, from 2006 to 2011, using participant responses (pathologist, cytotechnologist, and laboratory).DesignConcordance rates for the target diagnosis and general category for each slide challenge were analyzed. Four main factors were included in the analysis: (1) general category or specific responses, (2) program year from 2006 to 2011, (3) participant type (pathologist, cytotechnologist, or overall laboratory), and (4) preparation type (liquid-based or conventional).ResultsParticipants most frequently correctly classified negative for intraepithelial lesion or malignancy, low-grade squamous intraepithelial lesion, and herpes simplex virus infection, with concordance rates of 78.8%, 85%, and 80.2%, respectively. Performance on challenges with target diagnoses of high-grade squamous intraepithelial lesion (HSIL), squamous cell carcinoma, and ameba was poor, with concordance rates of 57.1%, 56.2%, and 41.5%, respectively. Significant improvement during the 6 years was seen in the concordance rates of participants' responses for low-grade squamous intraepithelial lesion challenges but not for HSIL. There was no significant difference in performance by slide preparation type.ConclusionsThe poor performance on anal cytology in the CAP NGC program, especially with regard to correct identification of HSIL and squamous cell carcinoma, indicates that there is a need for continued education about anal cytology.

ContextThe College of American Pathologists' Interlaboratory Comparison Program in Gynecologic Cytology has seen an increase in enrollment in liquid-based Papanicolaou test challenges with a decrease for conventional Papanicolaou tests. Trichomonas vaginalis can be difficult to identify in all preparation types.ObjectivesTo evaluate 20 years of participant results from the College of American Pathologists Interlaboratory Comparison Program in Gynecologic Cytology for Trichomonas to ascertain whether performance has changed because of the introduction of liquid-based Papanicolaou and proficiency testing.DesignConcordance rates for the target diagnosis of Trichomonas vaginalis were evaluated for 167,956 participant responses (1990-2010). A nonlinear mixed model was fit with participant type, preparation type, and a 2-level program year (1990-2005 and 2006-2010) reflecting before and after proficiency testing began. A repeated-measures component allowed modeling of the slide-specific performance to ensure that the overall results were not based on the performance of a few slides.ResultsCytotechnologists had higher concordance with the target diagnosis than did pathologists (89.8% [72,992 of 81,319] versus 83.4% [72,271 of 86,637], P < .001) and better performance for each preparation type (P = .003). Concordance initially dropped after the introduction of proficiency testing (P < .001) for conventional and liquid-based (SurePath) preparations by both participant types, followed by quick, parallel improvement.ConclusionsPerformance is high in the detection of Trichomonas vaginalis in the College of American Pathologists Interlaboratory Comparison Program in Gynecologic Cytology. Liquid-based Papanicolaou and proficiency testing minimally affected participant performance. Cytotechnologists performed better over time and across preparation types than did pathologists, although pathologists showed performance results parallel to that of the cytotechnologists. Awareness of the performance differences by pathologists and cytotechnologists, as well as their difference in proficiency among liquid-based techniques, may help ensure accurate results in clinical practice.

ContextAtrophic vaginitis is a commonly reported subset of Papanicolaou test results that are negative for intraepithelial lesion or malignancy, but interpretive criteria overlap with atrophic changes and other entities, hindering concordance among observers.ObjectivesTo report on the participant concordance from 2000 to 2009 in the College of American Pathologists Interlaboratory Comparison Program in Gynecologic Cytopathology, with a reference interpretation of atrophic vaginitis, and to investigate cytologic features of good and poorly performing slides to identify criteria useful in the interpretation of atrophic vaginitis.DesignWe summarized 18 302 responses from the program for slides with a reference interpretation of atrophic vaginitis. We randomly selected 18 Papanicolaou test results (3 conventional, 4 SurePath, and 11 ThinPrep) from good and poor performers for prospective, blinded criteria scoring for the following features: abundance of neutrophils, more than 100 degenerating parabasal cells, more than 25% necrotic background, more than 100 pseudoparakeratotic cells, and the presence of stripped or streaked nuclei, histiocytes, and superficial or intermediate squamous cells.ResultsMost Papanicolaou test results (>90%) with a specific reference interpretation of atrophic vaginitis were categorized as negative. Cytotechnologists are more likely than pathologists are to label it negative for intraepithelial lesion or malignancy (NILM) and are equally likely to mistake it for a high-grade lesion. Degenerating parabasal cells, pseudoparakeratosis, and necrotic background are associated with atrophic vaginitis (P  =  .001) on Papanicolaou. Abundant neutrophils (>100 per ×400 field) are also significantly correlated (P  =  .01).ConclusionsExact concordance to atrophic vaginitis is less than 90%. Most of the discrepancies are negative results for intraepithelial lesion or malignancy. Advanced atrophic features are as significant as neutrophils are to the interpretation of atrophic vaginitis.

Context.--Fluorescence in situ hybridization (FISH) and brightfield in situ hybridization (ISH) are 2 clinically approved laboratory methods for detecting ERBB2 (HER2) amplification in breast cancer. Objective.--To compare the performance of FISH and brightfield ISH on proficiency testing administered by the College of American Pathologists Laboratory Accreditation Program. Design.--Retrospective review was performed on 70 tissue core samples in 7 separate proficiency testing surveys conducted between 2009 and 2013. Results.--The samples included 13 consensus-amplified tissue cores, 53 consensus-nonamplified cores, and 4 cores that did not reach consensus for FISH and/or brightfield ISH. There were 2552 individual responses for FISH and 1871 individual responses for brightfield ISH. Consensus response rates were comparable for FISH (2474 of 2524; 98.0%) and brightfield ISH (2135 of 2189; 97.5%). The FISH analysis yielded an average HER2 copy number per cell that was significantly higher (by 2.86; P = .02) compared with brightfield ISH for amplified cores. For nonamplified cores, FISH yielded slightly, but not significantly, higher (by 0.17; P = .10) HER2 copy numbers per cell. There was no significant difference in the average HER2 to control ratio for either consensus-amplified or consensus-nonamplified cores. Participants reported 'unable to analyze' more frequently for brightfield ISH (244 of 2453; 9.9%) than they did for FISH (160 of 2684; 6.0%). Conclusions.--Our study indicates a high concordance rate in proficiency testing surveys, with some significant differences noted in the technical performance of these assays. In borderline cases, updated American Society of Clinical Oncology/College of American Pathologists cutoff thresholds that place greater emphasis on HER2 copy number per cell could accentuate those differences between FISH and brightfield ISH. doi: 10.5858/arpa.2017-0457-CP, Amplification or overexpression of the human epidermal growth factor receptor 2 (ERBB2, HER2) gene is an important oncogenic driver in up to 20% to 25% of breast cancers. Breast cancers [...]

Context.--Cerebrospinal fluid cytology is a critical diagnostic tool for the diagnosis of many conditions affecting the central nervous system. Objective.--To assess the performance characteristics of cerebrospinal fluid cytology samples by evaluating participant interpretations within the College of American Pathologists Nongynecologic Cytopathology Education program. Design.--Participant interpretations (N = 46 264) evaluated in the College of American Pathologists Nongynecologic Cytopathology Education Program were examined for concordance with the general category and with the reference diagnosis. Two nonlinear mixed models were used to analyze the concordance rates. Results.--The overall concordance rates for the general category and reference diagnosis were 92.1% and 81.0%, respectively. In the malignant category, the concordance rates with the reference diagnosis were lowest for diagnoses of nonhematopoietic small blue round cell tumors (54.8%) and metastatic malignancy (77.5%); the concordance rate with the reference diagnosis was highest for leukemia/ lymphoma (94.0%). In the benign category, the concordance rate was lowest for normal cerebrospinal fluid reference diagnoses (58.6%), followed by acute and chronic inflammation (64.6%), fungal infection (80.8%), and macrophages (85.3%). Significant differences in concordance were uncovered when performance was evaluated by participant type and stain technique. Leukemia/lymphoma was the most common diagnosis for misclassified nonhematopoietic small blue round cell tumor cases and negative or inflammatory cerebrospinal fluid cases. Conclusions.--This study illustrates the difficulties in achieving accurate diagnoses from cerebrospinal fluid specimens, particularly for nonhematopoietic small blue round cell tumors and normal and inflammatory cerebrospinal fluid specimens. (Arch Pathol Lab Med. 2018;142:833-837; doi: 10.5858/arpa.2017-0167-CP), Cerebrospinal fluid (CSF) examination is a well-established procedure for the evaluation and diagnosis of a wide variety of conditions affecting the central nervous system, including many cancers, infections, and inflammatory [...]

Context.--The College of American Pathologists (CAP) developed protocols for reporting pathologic characteristics of breast cancer specimens, including margin status. The Society of Surgical Oncology (SSO) and the American Society for Radiation Oncology (ASTRO) published treatment guidelines regarding margins in patients with invasive cancer; and SSO, ASTRO, and the American Society of Clinical Oncology (ASCO) recently published guidelines for patients with ductal carcinoma in situ. Objective.--To assess current practices among pathologists with regard to the processing/reporting of breast specimens, assess compliance with CAP cancer protocols, and assess alignment with SSO/ASTRO and SSO/ASTRO/ ASCO guidelines. Design.--A survey concerning breast specimen processing/reporting was distributed to pathologists enrolled in the CAP Performance Improvement Program in Surgical Pathology. Results.--Ninety-four percent (716 of 764 respondents) and 91% (699 of 769 respondents) define positive margins as 'tumor on ink' for invasive cancer and ductal carcinoma in situ, respectively, in compliance with CAP cancer protocols and with SSO/ASTRO and SSO/ASTRO/ ASCO guidelines. Of 791 respondents who provided details regarding methods for margin evaluation, 608 (77%) exclusively examine perpendicular margins, facilitating guideline compliance. However, 183 of 791 respondents (23%) examine en face margins in at least a subset of specimens, which may preclude guideline compliance in some cases. When separate cavity (shave) margins are examined, while 517 of 586 respondents (88%) ink these specimens, 69 of 586 (12%) do not, and this may also preclude guideline compliance in some cases. Conclusions.--A substantial proportion of survey participants report margin status for breast cancer specimens in a manner consistent with CAP cancer protocols, and in alignment with SSO/ASTRO and SSO/ASTRO/ASCO guidelines. However, there are opportunities for some laboratories to modify procedures in order to facilitate more complete adherence to guidelines. doi: 10.5858/arpa.2016-0626-CP, The College of American Pathologists (CAP) convened multidisciplinary consensus panels including physicians with expertise in breast cancer, who subsequently developed comprehensive protocols regarding the examination and reporting of surgical specimens [...]

Context.--Serum tests used for the screening and diagnosis of monoclonal gammopathies include serum protein electrophoresis (SPE; agarose gel or capillary zone), immunofixation (IFE) and immunosubtraction capillary electrophoresis, serum free light chains, quantitative immunoglobulins, and heavy/light-chain combinations. Urine protein electrophoresis and urine IFE may also be used to identify Bence-Jones proteinuria. Objective.--To assess current laboratory practice for monoclonal gammopathy testing. Design.--In April 2016, a voluntary questionnaire was distributed to 923 laboratories participating in a protein electrophoresis proficiency testing survey. Results.--Seven hundred seventy-four laboratories from 38 countries and regions completed the questionnaire (83.9% response rate; 774 of 923). The majority of participants (68.6%; 520 of 758) used agarose gel electrophoresis as their SPE method, whereas 31.4% (238 of 758) used capillary zone electrophoresis. The most common test approaches used in screening were SPE with reflex to IFE/immunosubtraction capillary electrophoresis (39.3%; 299 of 760); SPE only (19.1%; 145 of 760); SPE and IFE or immunosubtraction capillary electrophoresis (13.9%; 106 of 760); and SPE with IFE, serum free light chain, and quantitative immunoglobulins (11.8%; 90 of 760). Only 39.8% (305 of 767) of laboratories offered panel testing for ordering convenience. Although SPE was used by most laboratories in diagnosing new cases of myeloma, when laboratories reported the primary test used to follow patients with monoclonal gammopathy, only 55.7% (403 of 724) chose SPE, with the next most common selections being IFE (18.9%; 137 of 724), serum free light chain (11.7%; 85 of 724), and immunosubtraction capillary electrophoresis (2.1%; 15 of 724). Conclusions.--Ordering and testing practices for the screening and diagnosis of monoclonal gammopathy vary widely across laboratories. Improving utilization management and report content, as well as recognition and development of laboratory-directed testing guidelines, may serve to enhance the clinical value of testing. doi: 10.5858/arpa.2017-0128-CP, Monoclonal gammopathies are hematologic disorders characterized by abnormal production of one or more immunoglobulin clones. (1) They range from asymptomatic, benign disorders (such as monoclonal gammopathy of undetermined significance) to [...]

Context.--Body fluid cytology is an important diagnostic tool used to identify various conditions. However, an accurate diagnosis in this setting can sometimes be challenging.Objective.--To identify the performance characteristics of body fluid cytology by analyzing participant responses from the College of American Pathologists Interlaboratory Comparison Program in Nongynecologic Cytopathology.Design.--Participant responses from 5102 slides were analyzed for concordance to the general category (GC) and to the reference diagnosis (RD). Nonlinear mixed models were used to analyze concordance.Results.--The overall GC concordance was 95.2%. The GC type, participant type, and preparation type were significantly associated with GC concordance (P < .001). Concordance for malignant cases was higher than it was for benign cases. Cytotechnologists had better GC concordance compared to pathologists. ThinPrep (Hologic, Marlborough, Massachusetts) slides had the highest GC concordance. Participant type, fluid type, preparation type, and participant interpretation were significantly associated with rD concordance (P < .001). Pathologists performed better than cytotechnologists did for RD concordance. Pericardial fluid had the lowest RD concordance, especially for cases with normal or reactive findings. Modified Giemsa-stained slides performed best for lymphoma and hematopoietic malignancy. Small cell carcinoma had the highest GC concordance, and its RD concordance was higher in pleural than in peritoneal fluids. Adenocarcinoma showed the highest concordance rates for both GC and RD.Conclusions.--This study illustrates the challenges associated with interpreting body fluid cytology, particularly in pericardial fluid, and the factors that may affect accurate diagnoses. The results also highlight the value of using multiple preparation types in challenging cases.(Arch Pathol Lab Med. 2018;142:53-58; doi: 10.5858/arpa.2016-0509-CP), Body fluid cytology is an important diagnostic test for various malignant and benign conditions. Effusions can be caused by inflammatory, infectious, and benign; neoplastic or malignant; and primary or metastatic [...]

Context.--Laboratories must demonstrate analytic validity before any test can be used clinically, but studies have shown inconsistent practices in immunohistochemical assay validation. Objective.--To assess changes in immunohistochemistry analytic validation practices after publication of an evidence-based laboratory practice guideline. Design.--A survey on current immunohistochemistry assay validation practices and on the awareness and adoption of a recently published guideline was sent to subscribers enrolled in one of 3 relevant College of American Pathologists proficiency testing programs and to additional nonsubscribing laboratories that perform immunohistochemical testing. The results were compared with an earlier survey of validation practices. Results.--Analysis was based on responses from 1085 laboratories that perform immunohistochemical staining. Of 1057 responses, 65.4% (691) were aware of the guideline recommendations before this survey was sent and 79.9% (550 of 688) of those have already adopted some or all of the recommendations. Compared with the 2010 survey, a significant number of laboratories now have written validation procedures for both predictive and nonpredictive marker assays and specifications for the minimum numbers of cases needed for validation. There was also significant improvement in compliance with validation requirements, with 99% (100 of 102) having validated their most recently introduced predictive marker assay, compared with 74.9% (326 of 435) in 2010. The difficulty in finding validation cases for rare antigens and resource limitations were cited as the biggest challenges in implementing the guideline. Conclusions.--Dissemination of the 2014 evidencebased guideline validation practices had a positive impact on laboratory performance; some or all of the recommendations have been adopted by nearly 80% of respondents. (Arch Pathol Lab Med. 2017;141:1247-1254; doi: 10.5858/arpa.2016-0558-CP), The College of American Pathologists (CAP) Pathology and Laboratory Quality Center (the CAP Center) develops and maintains evidence-based laboratory practice guidelines (LPGs) that follow the US Institute of Medicine's Clinical [...]