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The authors questioned whether an automated kinetic mode assay of combined cytomegalovirus (CMV) late viral message and immediate and early antigens might result in a more sensitive and timely CMV diagnosis relevant to speedy treatment in the transplant setting. Toward this end, two cohorts were studied using automated in situ hybridization (ISH) for CMV as well as immunohistochemistry (IHC). The first cohort of patients consisted of 19 cases that were histologically positive (CMV-associated cytopathic change). A second cohort consisted of 10 cases that were histologically negative, yet culture positive. From the first cohort of histologically positive cases, 100% were positive by both ISH and IHC run on separate slides. In the second cohort, CMV was detected overall in 70% of cases (50% by ISH alone and 30% by IHC alone). These results indicate that a combined assay of ISH and IHC can detect more cases than routine hematoxylin and eosin staining or either assay alone. In two illustrative cases, used to demonstrate the feasibility of combining ISH and IHC, the authors used a combined two-color assay (ISH and IHC) performed sequentially on the same slide. The combined assays resulted in colocalized single cell message and protein in some cells and demonstrated more positive cells overall (some positive by IHC alone, some by ISH alone, and some by both) than either assay alone. The combined dual color assay can be completed within 4 to 5 hours giving the prospect of a same day result, which is faster than shell vial technique with immunofluorescence (24 to 48 hours) or culture (7 to 14 days). This study demonstrates that combining CMV message and protein assays results in a more sensitive assay and, when carried out in the kinetic mode, allows a speedy result relevant to early anti-CMV therapy.

As the demand for immunohistochemical tests (IHC) extends beyond artistry to more widespread medical diagnostic need, the field requires improvements in quality, reproducibility, speed, and standardization. Automated IHC offers the opportunity to impart new levels of quality, reproducibility, and standardization while reducing labor and reagent costs. In addition, the controlled environment of automated IHC may increase the speed and timeliness of results. Described is a computerized bar code-driven automatic immunostaining device that automatically dispenses reagents, controls washing, mixing, and heating to optimize IHC reaction kinetics, allowing assay within 60 minutes. This electronically controlled device generates high-quality, reproducible IHC assays in a medically timely manner.

A case of phagocytic, lambda light chain, plasma cell myeloma was characterized by its clinical, morphologic, cytochemical, immunologic, and cell kinetic features. A 40-year-old man presented with Coombs-negative hemolytic anemia, hepatosplenomegaly, lytic bone lesions, lambda light chain monoclonal gammopathy, and infiltration of the bone marrow by dysplastic plasma cells, 10% of which demonstrated phagocytosis of erythroid cells. Electron microscopy demonstrated myeloma cells with prominent cytoplasmic microfilaments and erythroid cells in intracytoplasmic vacuoles. The myeloma cells did not phagocytose staphylococci in vitro. Phagocytic and nonphagocytic myeloma cells were tartrate-sensitive, acid-phosphatase positive, alpha-napthyl butyrate esterase negative, and did not form E rosettes or EAox(IgG) rosettes. The tumor cells were Tdt, Ia antigen, and SIg negative. Immunofluorescent staining for cytoplasmic light chains showed a monoclonal lambda pattern in nonphagocytic myeloma cells, and a probable monoclonal lambda pattern in phagocytic myeloma cells. These findings characterize the neoplasm as a monoclonal proliferation of differentiated plasma cells with the capability of erythrophagocytosis. Erythrophagocytosis by myeloma cells may have been responsible for the hemolytic anemia. The tritiated thymidine labeling index (LI%) was high (8%), suggesting a poor prognosis, despite a dramatic initial response to chemotherapy.

Eleven patients with mature or peripheral T-cell lymphoma (PTL), other than mycosis fungoides, were identified using an extensive battery of T- and B-cell markers. Eight cases had a histologic diagnosis of either diffuse large cell or mixed lymphoma, three of small cell type. All cases had one or more "mature" T-antigens and an absence of B- and immature T-antigens. Assessment of T-antigens included E-rosettes (Er), anti-Leu 1-7 and Tdt. The authors delineated striking heterogeneity of T-antigen expression: 9 different immunotypes in 11 cases. Subset T-antigen assessment indicated T-helper neoplastic cells in five cases and T-suppressor in two. The remaining four had universal T-antigens alone. Seven cases appeared to have "novel" T-phenotypes not corresponding to normal T-ontogeny phenotypes. Novel or idiosyncratic phenotypes may be a key characteristic of PTL. Since no single T-antigen, including Er and Er receptor (Leu-5), was expressed in all cases, a battery of monoclonal antibodies is necessary to detect PTL. Clinically, the authors found PTL unexpectedly aggressive, despite mature immunotype. Most patients were elderly (median age 69); all had extranodal disease with cutaneous involvement (six cases) most frequent. Responses to chemotherapy frequently proved transient, with median survival of nine months. A fulminant course was noted even with localized presentation. Clinical outcome suggests PTL requires new therapeutic strategies.

To establish the feasibility of immunotyping specimens transported over great distance, the University of Arizona and the Cleveland Clinic exchanged 39 lymphoma cases (26 B-cell, 13 T-cell cases) via air express for phenotyping. They were exchanged with the use of glass slides with adherent snap-frozen sections. The method involved immunohistochemistry using a common battery of 20 antibodies directed at B- and T-cell antigens. Using similar, but not identical, methods and reagents, cases were assayed before and after transport. Results were generated as paired determinations in a double-blind fashion (1,338 total determinations, 669 paired determinations/669 antigens). Using "pretransport" values as the baseline, there was subsequent agreement with baseline in 624 of 669 antigenic determinations (93.3%) and disagreement in 45 (6.7%). Interinstitutional agreement was 94% for T-antigens (Leu-1-7,9) and 93% for B-antigens (K, lambda; IgM, G, A, D; B1, B2, B4, L14; J5, common acute lymphocyte leukemia antigen [CALLA]; Ia). In spite of 6.7% disagreement, there was agreement of overall phenotypic profile (e.g., B vs. T and subset stage) in all 39 cases. This is because discrepancies were isolated and the battery offset undue reliance on a single antigen. The disagreements relate mainly to certain B-antigens: CALLA/J5 (eight disagreements) and B1 and B2 (six and five disagreements, respectively). T-antigen disagreements were more sporadic (e.g., Leu-5 [three disagreements]). These discrepancies may relate to antigen loss with transport, variable antigen density or antibody avidity, methodologic differences, quality of reagents, or differences of phenotype interpretation. A further exchange of the 45 disagreement slides indicated that difference of observer interpretation was a minor factor (3 of 45). In conclusion, the authors' study demonstrates a high degree of immunotype reproducibility (approximately 93%), suggesting promise for institutions doing collaborative lymphoma studies.

Cytochemical, ultrastructural, and terminal deoxynucleotidyl transferase determinations in three cases of acute lymphocytic leukemia with azurophilic granules and punctate alpha-naphthyl acetate esterase (ANAE), alpha-naphthyl butyrate esterase (ANBE), and acid phosphatase (AP) activities further described a cytochemical and morphologic variant of acute lymphocytic leukemia. The ANAE, ANBE, and AP activities appeared specially localized within cytoplasmic granules. Electron microscopy substantiated a prominence of membrane-bound dense-core cytoplasmic granules. The variegated appearance of these dense-core granules and the cytochemical profile suggest that the aberrant granules are lysosomal in nature. Both the prominent cytoplasmic granules and the older age distribution led to early misdiagnosis of myelocytic leukemia. The distinctive cytochemical/morphologic profile and the ease of misdiagnosis suggest an important morphologic variant of acute lymphocytic leukemia.