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Abstract: A novel ultra‐low‐cost biochemical analysis platform to quantify protein dissociation binding constants and kinetics using paper microfluidics is reported. This approach marries video imaging with one of humankind's oldest materials: paper, requiring no large, expensive laboratory equipment, complex microfluidics or external power. Temporal measurements of nanoparticle–antibody conjugates binding on paper is found to follow the Langmuir Adsorption Model. This is exploited to measure a series of antibody–antigen dissociation constants on paper, showing excellent agreement with a gold‐standard benchtop interferometer. The concept is demonstrated with a camera and low‐end smartphone, 500‐fold cheaper than the reference method, and can be multiplexed to measure ten reactions in parallel. These findings will help to widen access to quantitative analytical biochemistry, for diverse applications spanning disease diagnostics, drug discovery, and environmental analysis in resource‐limited settings. [ABSTRACT FROM AUTHOR]

Objectives The immune trypanolysis test ( TL) is an accurate sero-diagnostic tool increasingly implemented for sleeping sickness medical surveillance, but it is restricted to the reference laboratories. To facilitate storage and transport of the test specimen, we developed a protocol for the examination of blood spotted on filter paper ( TL-fp) that can be stored and shipped at ambient temperature. We compared its performance with the classical TL on plasma ( TL-pl) that needs to be kept frozen until use. Methods The study was conducted in active foci of the Republic of Guinea. In total, 438 specimens from treated and untreated sleeping sickness patients and serological suspects were tested with both methods. Result TL-fp gave significantly less positive results than TL-pl, but all the confirmed sleeping sickness cases were positive with the TL-fp protocol. Conclusion TL-fp appears to offer a good compromise between feasibility and sensitivity to detect currently infected subjects who play a role in the transmission of Trypanosoma brucei gambiense and is useful for contributing to the elimination of gambiense sleeping sickness. [ABSTRACT FROM AUTHOR]

Proteins immobilized on insoluble carriers or immunoadsorbents are now widely used in many areas of biological research. They are especially efficient in studies of complex protein mixtures because immunoadsorbents are very useful for isolation of individual components for further investigation. Antibodies immobilized on paper retain their specific activity and can be applied as very sensitive measurement tool. [ABSTRACT FROM AUTHOR]

A combination of blotting of bacterial colonies on nitrocellulose paper discs and immunoenzymatic detection of bound antigens with specific monoclonal antibodies was used to detect and serotype Actinobacillus actinomycetemcomitans directly off the initial agar culture dish. The A. actinomycetemcomitans antigens, representative of specific colonies, were identified immunoenzymatically using monoclonal antibody specific for a species-specific antigen. The serotype of the bacteria in colonies was identified by dividing the blotting paper into sections and immunoenzymatically identifying the serotype antigens with serotype-specific monoclonal antibodies. This procedure provides for simple, rapid, sensitive and accurate identification and characterization of A. actinomycetemcomitans isolates from the initial isolation agar plates. [ABSTRACT FROM AUTHOR]

Most Human African Trypanosomiasis (HAT) control programmes in areas endemic for Trypanosoma brucei gambiense rely on a strategy of active mass screening with the Card Agglutination Test for Trypanosomiasis (CATT)/T. b. gambiense. We evaluated the performance, stability and reproducibility of the CATT/T. b. gambiense on blood-impregnated filter papers (CATT-FP) in Kajo-Keji County, South-Sudan, where some areas are inaccessible to mobile teams. The CATT-FP was performed with a group of 100 people with a positive CATT on whole blood including 17 confirmed HAT patients and the results were compared with the CATT on plasma (CATT-P). The CATT-FP was repeated on impregnated filter papers stored at ambient and refrigerated temperature for 1, 3, 7 and 14 days. Another 82 patients with HAT, including 78 with a positive parasitology, were tested with the CATT-FP and duplicate filter paper samples were sent to a reference laboratory to assess reproducibility. The CATT-FP was positive in 90 of 99 patients with HAT (sensitivity: 91%). It was less sensitive than the CATT-P (mean dilution difference: -2.5). There was no significant loss of sensitivity after storage for up to 14 days both at ambient and cool temperature. Reproducibility of the CATT-FP was found to be excellent (kappa: 0.84). The CATT-FP can therefore be recommended as a screening test for HAT in areas where the use of CATT-P is not possible. Further studies on larger population samples in different endemic foci are still needed before the CATT-FP can be recommended for universal use. [ABSTRACT FROM AUTHOR]

We reviewed eight cases that were diagnosed before 1995 with B-prolymphocytic leukaemia (B-PLL) harbouring t(11;14)(q13;q32) and/or cyclin D1 staining. Thirteen B-PLL patients without t(11;14) were selected as controls. Peripheral blood, bone marrow and histological sections were re-examined without cytogenetic information. Final diagnosis was made using morphology, cytogenetics, immunophenotype and immunohistochemistry. Clinical characteristics were similar for both groups except for younger age, male predominance and extranodal involvement in cases with t(11;14). CD5 was more frequently positive in the t(11;14)+ group (80%) than in the t(11;14)− group (31%). Surface membrane immunoglobulin was strongly expressed by all t(11;14)+ cases, but only 45% of t(11;14)− cases. Histopathological and cytological review of cases with t(11;14) showed an infiltrate with a mixture of cells, some resembling prolymphocytes and others with mantle cell lymphoma (MCL) morphology. Blood films of cases with t(11;14) showed features suggestive of B-PLL in three, and in others, a mixture of cells resembling MCL and nucleolated ones; none corresponded to the blastoid form of MCL. We suggest that ‘B-PLL’ with t(11;14) may represent a splenomegalic form of MCL evolving with leukaemia. These cases illustrate the importance of tissue diagnosis with cyclin D1 staining and fluorescence in situ hybridization analysis in B-cell leukaemia with prolymphocytic features. [ABSTRACT FROM AUTHOR]

Because of its B-cell depleting effect, rituximab has entered clinical trials in several autoimmune conditions. This study assesses the efficacy and safety of rituximab in 57 adults with chronic immune thrombocytopenic purpura (ITP). All patients had platelet counts <30 × 109/l, all had received two or more previous ITP treatments and 31 had undergone splenectomy. Patients received rituximab 375 mg/m2 weekly for 4 weeks. Thirty-one patients (54%) responded, achieving a platelet count >50 × 109/l: 18 achieved a complete response (CR: platelet count >150 × 109/l) and 13 a partial response (PR: platelet count 50–150 × 109/l). Twenty-nine responses occurred within 8 weeks of the first infusion. Sixteen of 18 CR patients (28% overall), including eight who had failed splenectomy, continued in CR after a median of 72·5 weeks; 15 of 16 are >1 year from the first infusion. Only two of 13 maintained a PR. Thirty-three patients experienced grade 1–2 adverse events and one a grade 3 event, but they all completed treatment. Circulating B cells fell to <0·03 × 109/l. No changes in immunoglobulin levels or infectious complications were seen. In summary, rituximab was well tolerated with no immediate complications and induced a lasting, substantial response in 32% of adults with chronic ITP. [ABSTRACT FROM AUTHOR]

The frequency and prognostic relevance of translocations t(11;14) and t(4;14), the most common translocations involving the immunoglobulin heavy chain (IgH) gene in multiple myeloma (MM), were investigated in 128 patients treated with intensive chemotherapy and autologous stem cell transplant. Myeloma cells were identified by cytoplasmic light chain immunofluorescence combined with fluorescence in situ hybridization (cIg-FISH) for detection of translocations t(11;14) and t(4;14). Overall, t(11;14) was detected in 16 of 125 (12·8%) and t(4;14) in 15 of 120 (12·5%) patients. Progression-free and overall survivals were similar for patients with or without t(11;14). However, patients with t(4;14) had significantly shorter progression-free (median 9·9 months vs. 25·8 months; P = 0·0003) and overall survivals (median 18·3 months vs. 48·1 months; P < 0·0001) than patients without t(4;14). The t(4;14) was associated with IgA and t(11;14) with light chain MM. There was no association between the t(11;14) or t(4;14) and other biological parameters including age, gender, haemoglobin, β-2 microglobulin, C-reactive protein, calcium, creatinine, albumin, or the percentage of bone marrow plasma cells. Multivariate analysis identified t(4;14) as the only adverse prognostic factor for both progression-free survival and overall survival. Our results indicate that the t(4;14) detected by cIg-FISH is associated with a poor prognosis in MM patients receiving intensive chemotherapy and autotransplant. [ABSTRACT FROM AUTHOR]

Single nucleotide polymorphisms (SNPs) of inflammatory cytokine genes were examined in 84 adult Japanese patients with chronic immune thrombocytopenic purpura (ITP) and 56 race-matched healthy controls. The SNPs examined were within the genes encoding tumour necrosis factor (TNF)- α (−238 G/A and −308 G/A), TNF- β (+252 G/A), and interleukin (IL)-1 β (−511 C/T and +3953 T/C). Of these SNPs, the frequency of the TNF- β (+252) G/G phenotype was significantly higher in ITP patients than in healthy controls (21% vs. 7%, P = 0·04, odds ratio = 3·6, 95% confidence interval 1·1–11·1), while no significant association was detected for the other SNPs. The distribution of the TNF- β (+252) phenotype was not associated with human leucocyte antigen class II alleles or the therapeutic response in ITP patients. The frequency of circulating anti-glycoprotein IIb/IIIa antibody-producing B cells was significantly higher in ITP patients with the TNF- β (+252) G/G phenotype than in those with the G/A or A/A phenotype (11·9 ± 4·9 vs. 6·8 ± 4·9 and 3·7 ± 2·8 per 105 peripheral blood mononuclear cells; P = 0·02 and P < 0·001, respectively). These findings suggest that the SNP located at TNF- β (+252) contributes to susceptibility to chronic ITP by controlling the autoreactive B-cell responses to platelet membrane glycoproteins. [ABSTRACT FROM AUTHOR]

We have investigated whether intracellular signal transduction molecules can be used as immunohistological markers of normal and neoplastic human leucocytes in routine tissue sections. We obtained selective labelling of white cells for eight such molecules (the ‘linker’ molecules SLP-76 and BLNK, the Src family kinases Lyn, Fyn, Syk and Hck, and the phospholipases PLC- γ1 and PLC- γ2). Antibodies to SLP-76 and PLC- γ1 selectively labelled T cells, and antibodies to BLNK, Lyn, Fyn, Syk and PLC- γ2 labelled B cells (although Fyn immunostaining was restricted to mantle zone B cells). Antibodies to the Syk and Hck kinases labelled probable thymocyte precursors at the periphery of the thymic cortex. In addition to lymphoid cells, several other leucocyte types were immunostained (e.g. SLP-76, Lyn, Syk and Hck were found in megakaryocytes, myeloid cells and/or macrophages, and PLC- γ2 was detected in arterial endothelium). SLP-76 and PLC- γ1 were found in most T-cell lymphomas studied, and some B-cell lymphomas were also positive for PLC- γ1 (e.g. diffuse large cell and Burkitt's lymphoma). The five B cell-associated markers were found in most B-cell non-Hodgkin's lymphomas, although some diffuse large B-cell lymphomas were negative (e.g. for Lyn) and anti-Fyn tended not to stain small B-cell neoplasms. The observation that a range of leucocyte signalling molecules can be detected in routine biopsies offers new possibilities for studying normal and neoplastic human white cells in diagnostic tissue samples. [ABSTRACT FROM AUTHOR]

Intravenous (IV) anti-D and IV immunoglobulin (IVIG) slow the Fc γ receptor (Fc γR)-mediated destruction of antibody-coated platelets in patients with immune thrombocytopenic purpura (ITP). This pilot study explored the mechanism of these immunoglobulin preparations by measuring interleukin-10 (IL-10), monocyte chemoattractant protein-1 (MCP-1), IL-6 and tumour necrosis factor  α (TNF α), before and after infusion and by assessing the effect of Fc γRIIa and Fc γRIIIa polymorphisms on both cytokine and haematologic responses to anti-D. Following IVIG, only IL-10 was increased at 2 h and MCP-1 on day 7 ( P < 0·05). In contrast, 2 h after anti-D infusion, plasma levels of all four cytokines were increased ( P < 0·01); five of six patients with the highest MCP-1, IL-6 and TNF α levels had chills. Higher IL-10 levels correlated with platelet increases at 24 h and haemoglobin decreases at day 7 ( P < 0·025). Patients with the Fc γRIIa-131HH genotype had significantly higher MCP-1, IL-6 and TNF α levels. Patients with the Fc γRIIIa-158VF genotype had higher platelet increments at day 7 ( P < 0·05). Soluble CD16 (sCD16) was increased 2 h after IV anti-D; day 7 levels correlated with day 7 haemoglobin decreases ( P < 0·01). In conclusion, the relationship of Fc γRIIa and Fc γRIIIa polymorphisms with both cytokine levels and platelet increments implicated these receptors in responses to anti-D and supported different mechanisms of Fc γR interaction to those seen with IVIG. [ABSTRACT FROM AUTHOR]

This study was undertaken to investigate the prevalence of basement membrane zone (BMZ) antibodies, their subtypes and clinical correlations in 96 patients attending the Oxford vulval clinic with lichen sclerosus (LS) of the vulva. Indirect immunofluorescence of serum (intact and split skin) to immunoglobulin (Ig)G was performed looking for the presence or absence of staining at the BMZ. Eighteen patients' sera (14 with positive indirect immunofluorescence to IgG) were examined for IgG antibodies of subclasses IgG1, 2 and 3, and 23 sera were examined for IgG4 subclass. Immunoblotting was performed in seven patients, and showed antibodies to BP180 in six patients and BP230 in one. One-third of patients with vulval LS had BMZ antibodies binding to the epidermal side of salt split skin. Immunoblotting showed antibodies to BP180 collagen XVII (six of seven patients) and BP230 in one. The subclasses were chiefly IgG1 and 2, different from those seen in bullous pemphigoid. No clinical correlation was found between the presence of antibodies and the presence of erosions, severity of scarring, age of onset of disease or response to treatment. These antibodies may be a reflection of a tendency to produce autoantibodies or be relevant to pathogenesis. [ABSTRACT FROM AUTHOR]

Individuals with primary or secondary immune-deficiency diseases may be at risk for vaccinia infection if widespread smallpox-immunization programmes are implemented in the United States of America (USA) for bioterrorism preparedness. The objective of this study was to determine whether commercial immune globulin (intravenous, human) products contain biologically active antibodies to vaccinia that have the potential to protect people, with immune deficiencies, from complications of vaccinia. Eight currently United States (US)-licensed and two European intravenous immunoglobulin (IVIG) products were tested in a vaccinia plaque-reduction neutralization assay. The in vivo activity of five of these lots was assessed in severely immune-deficient mice. All tested products contained neutralizing anti-vaccinia activity, in vitro and in vivo. The use of IVIG by individuals with inherited or acquired humoral immune deficiencies may provide some protection if they are inadvertently exposed to vaccinia. [ABSTRACT FROM AUTHOR]

In cold agglutinin disease, monoclonal red blood cell autoantibodies, termed cold agglutinins, induce haemolysis in patients exposed to the cold. Commonly, these autoantibodies are directed against the developmentally regulated I/i blood groups. A second blood group system, the Pr system (located on glycophorins), is involved less frequently. Anti-Pr cold agglutinins recognize either alpha 2,3- or alpha 2,6-linked N-acetylneuraminic acid as the immunodominant group. Cold agglutinins of anti-I/i specificity show a remarkable restriction in their genomic repertoire of the immunoglobulin heavy and light-chain immunoglobulin-variable domain (i.e. exclusive use of VH4-34 in heavy chains). For anti-Pr cold agglutinins, preliminary data on the repertoire of the light-chain variable domain indicate a preference for the subgroup Vkappa IV. To elucidate restrictions in the light-chain variable-domain subgroup repertoire of anti-Pr cold agglutinins systematically, and to discuss these results in the context of their anti-Pr1–3 subclassification and immunodominant sialic acid, light chains in 13 anti-Pr cold agglutinins were investigated. The anti-Pr light chains were isolated using temperature-dependent absorption/elution techniques. Subsequently, they were subjected to N-terminal Edman degradation, and the light chain Vkappa subgroup was affiliated using the Kabat database. Five of 13 (38%) light chains belonged to Vkappa IV, five of 13 (38%) to Vkappa I and three of 13 (23%) to Vkappa III. Anti-Pr with Vkappa IV subgroup light chains exclusively recognized alpha 2,3-linked N-acetylneuraminic acid. Including data from the literature, the repertoire of the light-chain variable domain in pathological anti-Pr cold agglutinins exhibits a clear bias towards the use of the single germline gene-derived subgroup, Vkappa IV (eight of 17 or 47%). The association of Vkappa IV subgroup light chain-containing anti-Pr cold agglutinins with binding to alpha 2,3-, but not alpha 2,6-linked N-acetyneuraminic acid raises speculations about a possible role of subgroup-derived determinants in anti-Pr binding. [ABSTRACT FROM AUTHOR]

Mantle cell lymphoma (MCL) shares immunophenotypic and karyotypic features with chronic lymphocytic leukaemia. The latter comprises two distinct entities with prognosis dependent upon immunoglobulin heavy chain (IgH) gene mutational status and the presence of 11q deletion. We evaluated the relevance of IgH gene mutational status, IgV gene family usage and presence of 11q deletion in a series of 42 histologically reviewed classical MCL cases to determine the prognostic impact. VH3 was the most common VH family, with VH3-21 being the most frequent individual VH gene. Approximately 30% of the cases had a IgH somatic mutation rate higher than 2%, but was only higher than 4% in <10% of cases. Half of the cases had deletion of chromosome 11q21-telomere (11q21->ter), with two minimal deleted regions, at 11q22.2 and 11q23.2. There was no association between 11q loss and IgH gene somatic mutation rate; the use of VH3-21 gene could be associated with a better prognosis. [ABSTRACT FROM AUTHOR]

Systemic AL amyloidosis (AL) is a disorder in which light chains form fibrillar deposits, leading to organ dysfunction and death. Rarely, AL has been associated with non-Hodgkin's lymphoma (NHL), although this association has not been well characterized. We report a series of six patients with AL associated with NHL, primarily lymphoplasmacytic lymphoma. Organ involvement was variable, with frequent bulky lymphadenopathy and visceral cavity deposits, but no cardiac involvement. Positron emission tomography scans were negative. Bone marrow and lymph node biopsies showed a mixed population of CD20+ lymphoid and CD138+ plasma cells. Serum free light chains were elevated, and correlated with response to therapy. Immunoglobulin light chain variable region (Ig VL) germline gene use was typical for AL, reflecting previously observed correlations between germline gene use and organ tropism. Five patients received rituximab-based therapies with two responses. Two patients underwent autologous stem cell transplantation with one complete haematological response. Four patients survive at 10–132 months from diagnosis. AL with NHL has distinctive clinical features but employs the same Ig VL gene repertoire as AL with clonal plasma cell dyscrasias. Serial serum free light chain levels are useful for tracking response to therapy. Treatments aimed at both lymphoid and plasma cell components appear warranted. [ABSTRACT FROM AUTHOR]

Alloantibody tests demonstrate immunological causes of insufficient increments in random platelet transfusions. The value of a positive or negative test result in predicting the outcome of human leucocyte antigen (HLA)-matched transfusions in patients refractory to leucodepleted random platelet transfusions has not been assessed. We retrospectively evaluated the outcome of the first HLA-matched platelet transfusion in 72 patients with haematological diseases in two ways: first, the strategy according to which the patient was selected for HLA-matched platelet transfusions was analysed. The strategies were: (i) results of alloantibody tests were not available, (ii) a positive alloantibody test, (iii) a negative alloantibody test. Secondly, the outcome of the first HLA-matched transfusion was investigated relative to the results of alloantibody tests, irrespective of the decision strategy. No significant association was found between the decision strategy and the outcome of the first HLA-matched platelet transfusion. Positive alloantibody tests, however, predicted a better outcome of the first HLA-matched platelet transfusion ( P = 0·04 and P = 0·03 after 1 and 16 h respectively). In patients refractory to random platelet transfusions, positive alloantibody tests predicted a better outcome of HLA-matched platelet transfusions. Patients with negative alloantibody tests, however, may benefit from HLA-matched platelet transfusions. [ABSTRACT FROM AUTHOR]

This study was conducted to analyse the usefulness of hepatitis C virus (HCV) core antigen tests for the confirmation of HCV infection in a donor presenting as nucleic acid amplification technology (NAT) positive but negative for antibodies to HCV (anti-HCV). Blood donations were screened, in parallel, for anti-HCV using the Abbott PRISM HCV Chemiluminescent immunoassay (ChLIA) and an ‘in-house’ HCV NAT (pools of up to 95 donations). An HCV NAT-positive antibody-negative donor was identified. Twelve follow-up samples were obtained and tested with various HCV antigen (including the recently marketed Trak-C second-generation assay) and HCV antibody assays. The single HCV NAT-positive, antibody-negative donation was identified from 1 117 681 donations screened in the 4-year period, July 1999 to June 2003. The index donation was positive by Ortho HCV core antigen enzyme immunoassay (EIA) and Ortho Trak-C (second-generation HCV core antigen EIA). An archive sample, taken 127 days prior to the index donation, was negative for all HCV markers. Subsequent samples demonstrated a loss of reactivity in the Ortho HCV core antigen EIA and reduced activity in the Ortho Trak-C until day 69. Immunoblot (Ortho RIBA-3) and HCV PRISM became positive on day 62, whilst Ortho HCV ELISA was not positive until day 132 or Biorad HCV ELISA until day 160. An alternative immunoblot (Innogenetics Innolia III) was positive from day 55. RNA levels fluctuated considerably during the follow-up period, being completely undetectable by routine screening methods at the time-point around seroconversion; subsequently, antibody was detected using all assays investigated. This HCV-converting blood donor provided a unique panel of samples for using to assess current (and future) HCV assay systems. The overall test results led to the conclusion that individual HCV antigen testing should not be considered as equivalent to HCV NAT minipool screening. Trak-C antigen testing may be considered as a suitable confirmatory assay for isolated HCV NAT reactivity. [ABSTRACT FROM AUTHOR]

Administration of diclofenac may lead to immune haemolytic anaemia (IHA) owing to the presence of drug-dependent antibodies and/or autoantibodies. A relationship with oral or intramuscular drug administration is unknown. Here, we describe a patient who apparently tolerated oral diclofenac but developed severe IHA following intramuscular injection of the drug. A 66-year-old-female was admitted to hospital because of jaundice and nausea, which were initially presumed to be manifestations of a postcholecystectomy syndrome. The patient soon developed haemolysis and renal failure. Although the symptoms and signs were suggestive of autoimmune haemolytic anaemia (AIHA), the patient had diclofenac-induced IHA. Serological testing, including detection of drug-dependent antibodies, was performed using standard techniques. The patient's serum was found to contain a highly reactive diclofenac-dependent red cell antibody of the immune complex type (titre 256 000). She recovered after 7 weeks of treatment with prednisolone, blood transfusions, haemodialysis and plasma exchange. Diclofenac-induced IHA should always be considered when a patient on diclofenac develops haemolysis. [ABSTRACT FROM AUTHOR]

Objectives: Immunoglobulin A dominates mucosal surfaces and is a biomarker of interest in populations with a high disease burden. The objectives of this work are to describe an ELISA for IgA and test the stability of storage on filter paper for human milk and saliva collection to be used in remote field locations. Methods: A two-site sandwich ELISA for IgA was developed. To test filter paper storage capabilities under field conditions, 248 matched whole and dried human milk filter paper samples and 251 matched whole and dried saliva samples were collected from northern Kenyan women. Whole samples were frozen in liquid nitrogen while dried samples were stored at ambient temperature for up to 8 weeks. Recovered dried IgA levels were compared to whole IgA levels and adjusted for time stored at ambient temperature. Results: The lower limit of quantification for this assay is 10.1 ng/ml. Linearity of dilution for human milk and saliva samples was excellent. High and low-control coefficient of variation values across plates were 9.1 (341.8 ng/ml) and 9.4% (132.5 ng/ml). IgA was detected in all whole and dried samples. There is a moderate concordance between dried and whole samples ( R2 = 0.62). There is a small but significant effect of time stored, with a loss of ∼1 μg/ml per day ( P = 0.0052). Conclusions: This IgA assay is a cost-effective alternative to commercial secretory IgA kits. Human milk and saliva can be stored on filter paper for up to 8 weeks. Am. J. Hum. Biol., 2011. © 2011 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Detecting antibodies in blood is as simple as it can get. In their Communication on page 15369 ff., D. Citterio, M. Merkx et al. integrate antibody‐targeting bioluminescent sensing proteins and other essential assay components into a microfluidic paper‐based analytical device. A drop of blood, a digital camera, and twenty minutes are all that is required to detect the presence and the concentration of multiple antibodies in whole blood based on the color of the emitted light. [ABSTRACT FROM AUTHOR]

Lambert–Eaton myasthenic syndrome (LEMS) is a rare paraneoplastic neurological syndrome of the neuromuscular transmission. The symptoms often progress slowly and can be misdiagnosed in early stage. Seropositive SOX‐1 antibodies are support for the diagnosis of LEMS and have high specificity for small cell lung cancer (SCLC). In this paper, we report a case of a 56‐year‐old man with smoking history who was admitted to hospital with progressive muscle weakness of the proximal legs. LEMS was diagnosed by repetitive nerve stimulation (RNS) testing and seropositive SOX‐1 antibodies. Primary screening with chest computed tomography (CT) and integrated PET/CT did not reveal any tumor. After continuous follow‐up, SCLC was found by chest CT and confirmed with pathological examination 10 months after the diagnosis of LEMS. Long‐term follow‐up and screening for occult SCLC in LEMS patients with positive SOX‐1 antibodies are very important. [ABSTRACT FROM AUTHOR]

‘Natural’ opsonins to Salmonella typhimurium in serum from foetal and natal rats have been shown to be in part macroglobulins. The titre of these opsonins was shown to increase in the young natal animals. [ABSTRACT FROM AUTHOR]

Accurate surveillance of coronavirus disease 2019 (COVID‐19) incidence includes large‐scale antibody testing of the population. Current testing methods require collection of venous blood samples by a healthcare worker, or dried blood spot (DBS) collection using finger prick, however this might have some logistic and processing limitations. We investigated the performance of the Ser‐Col device for detecting severe acute respiratory syndrome coronavirus‐2 (SARS‐CoV‐2) antibodies using a finger prick: DBS‐like collection system that includes a lateral flow paper for serum separation and allows for automated large scale analysis. For this prospective study, adult patients with moderate to severe COVID‐19 were included 6 weeks post‐symptom onset. Healthy, adult volunteers were included as a negative control group. Venous blood and capillary blood using the Ser‐Col device were collected and the Wantai SARS‐CoV‐2 total antibody ELISA was performed on all samples. We included 50 subjects in the study population and 49 in the control group. Results obtained with venous blood versus Ser‐Col capillary blood showed 100% sensitivity (95% CI: 0.93−1.00) and 100% specificity (95% CI: 0.93−1.00). Our study shows the feasibility of SARS‐CoV‐2 total antibody screening using a standardized DBS technique with semiautomated processing for large scale analysis. [ABSTRACT FROM AUTHOR]

This article focuses on a list of forthcoming papers published in the journal. Some of them are: "Conformation and Domain Structure of the Non-Histone Chromosomal Proteins," "Differential Response of Cultured Parsley Cells to Elicitors from Two Non-Pathogenic Strains of Fungi," "Monoclonal Antibodies Against Platelet Membrane Glycoproteins," "Synthesis of Biotin-labelled Dexamethasone Derivatives: Novel Harmone-Affinity Probes," "On the Mechanisms of ATP-induced and Succinate-Induced Redistribution of Cations in Isolated Rat Liver Cells," "Shedding of Gangliosides From Calf Thymocytes."

The article provides updates on several veterinary research. Tatsuyuki Okinaga and colleagues used commercial enzyme-linked immunosorbent assay to test pig sera for antibodies to porcine reproductive and respiratory syndrome virus. Wojciech Zygner and colleagues described the results of testing blood samples from dogs for a range of tickborne pathogens. Ana Balseiro and colleagues reported cases of tuberculosis identified at postmortem examination of individual roe deer in Spain and Italy.

Presents several developments in the veterinary medicine. Identification of keel and furculum damage in laying hens; Detection of antigens in fresh and autolysed tissue; Hypervaccination with marker vaccines against bovine herpesvirus type 1.

We introduce a general method to transform antibodies into ratiometric, bioluminescent sensor proteins for the no-wash quantification of analytes. Our approach is based on the genetic fusion of antibody fragments to NanoLuc luciferase and SNAP-tag, the latter being labeled with a synthetic fluorescent competitor of the antigen. Binding of the antigen, here synthetic drugs, by the sensor displaces the tethered fluorescent competitor from the antibody and disrupts bioluminescent resonance energy transfer (BRET) between the luciferase and fluorophore. The semisynthetic sensors display a tunable response range (submicromolar to submillimolar) and large dynamic range (Δ Rmax>500 %), and they permit the quantification of analytes through spotting of the samples onto paper followed by analysis with a digital camera. [ABSTRACT FROM AUTHOR]

In this case series of four paediatric patients, we present the first described cases of immunotherapy‐responsive autoimmune nodopathy with IgG2 antineurofascin antibodies. In three cases, the antineurofascin antibodies were predominantly of the IgG2 subclass, a novel finding in comparison to previously described adult cases where IgG4 and/or IgG1/3 have typically been described. One patient had low signal for IgG2 with predominant IgG1 and IgG4 antibodies, a pattern commonly seen in adult patients. Two patients had antibodies targeting all three neurofascin isoforms (155, 186, and 140), whereas antibodies in the sera from the third targeted only the nodal isoforms 186 and 140, and the fourth patient only neurofascin 155. The three patients with IgG2 predominant antibodies appear to be responsive to intravenous immunoglobulin (IVIG) to varying degrees thus far, whereas the patient with IgG1/4 antibodies had poor response to IVIG but good response to steroids. Although the full clinical significance of IgG2 predominant antineurofascin antibodies in the context of childhood polyneuropathy remains unclear, emerging evidence of serological‐phenotypic correlation may inform prognostication and therapeutic decision‐making, warranting further study into this area. What this paper adds: Paediatric immunotherapy‐responsive nodopathies were associated with antineurofascin antibodies predominantly of the IgG2 subclass in 3 out of 4 patients.Identification of antibodies and understanding their phenotypic relevance could predict response to treatment and guide therapeutic decision‐making in children. [ABSTRACT FROM AUTHOR]

Symptoms of pelvic masses, elevated serum CA125 levels, massive ascites, and pleural effusion in female patients are usually associated with malignancy. Some benign ovarian tumors or other nonmalignant tumors may also produce similar symptoms, called Meigs syndrome or pseudo‐Meigs' syndrome, which should be one of the differential diagnoses. However, there is an extremely rare form of SLE called pseudo‐pseudo Meigs syndrome (PPMS), which may also present with the above symptoms, but is not associated with any of the tumors. In this paper, we report a case of a 47‐year‐old woman who presented with abdominal distention. The patient was found to have elevated serum CA125 levels to 182.9 U/mL before the operation. Her PET‐CT suggested a large heterogeneous mass in the pelvis measuring 8.2 × 5.8 cm with a large amount of ascites. She was initially diagnosed with ovarian cancer and underwent exploratory laparotomy. Pathology of the surgical specimen revealed a uterine leiomyoma. Two months after discharge, the patient's ascites reappeared along with recurrent intestinal obstruction. After ascites and serological tests, she was eventually diagnosed with systemic lupus erythematosus and received systemic hormonal therapy. [ABSTRACT FROM AUTHOR]

Globally, the eradication of bovine viral diarrhoea virus (BVDV) is still in its infancy, but eradication has been, or is being, adopted by several countries or regions. Comparisons between countries' schemes allow others to assess best practice, and aggregating published results from eradication schemes provides greater statistical power when analysing data. Aggregating data requires that results derived from different testing schemes be calibrated against one another. The authors aimed to evaluate whether relationships between published BVDV test results could be created and present the outcome of a systematic literature review following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. The results are tabulated, providing a summary of papers where there is potential cross-calibration and a summary of the obstacles preventing such data aggregation. Although differences in measuring BVDV present barriers to academic progress, they may also affect progress within individual eradication schemes. The authors examined the time taken to retest following an initial antibody BVDV test in the Scottish eradication scheme. The authors demonstrate that retesting occurred quicker if the initial not negative test was from blood rather than milk samples. Such differences in the response of farmers/veterinarians to tests may be of interest to the design of future schemes. [ABSTRACT FROM AUTHOR]

Sakiyama Y, Kato R, Inoue S, Suzuki K, Itabe H, Yamamoto M. Detection of oxidized low-density lipoproteins in gingival crevicular fluid from dental patients. J Periodont Res 2009; doi: 10.1111/j.1600-0765.2009.01226.x. © 2009 John Wiley & Sons A/S Background and Objective: Oxidative modification of low-density lipoprotein (LDL) occurs in various diseased tissues and sites of local inflammation. For example, an increased plasma oxidized low-density lipoprotein (OxLDL) level is a well-known risk marker for cardiovascular diseases. Gingival crevicular fluid, the exudate from gingival tissues into the sulci, can be easily collected in a non-invasive manner. However, the possible presence of OxLDL in gingival crevicular fluid has not been studied. In this study, we established a procedure to measure OxLDL in human gingival crevicular fluid. Material and Methods: Human gingival crevicular fluid was sampled with paper points or paper strips. The gingival crevicular fluid samples from healthy gingival sulci (pocket depth < 4 mm, n = 14) were subjected to western blot and/or sandwich ELISA. The amounts of OxLDL and LDL were measured by sandwich ELISA using an anti-oxidized phosphatidylcholine monoclonal antibody and two anti-apolipoprotein B antibodies. Venous blood samples were analyzed biochemically. Results:  We tested two methods of gingival crevicular fluid collection, namely paper points and paper strips. Gingival crevicular fluid could be collected very safely with paper points and they showed good recovery of LDL and OxLDL throughout the analysis. Apolipoprotein B, the major protein component in LDL, was detected in gingival crevicular fluid by western blot, and OxLDL was found to be present in gingival crevicular fluid by ELISA. The OxLDL/LDL ratio in gingival crevicular fluid was 17.0 times higher than that in plasma. Conclusion: This is the first report to show the presence of apolipoprotein B and apolipoprotein B– oxidized phosphatidylcholine complex, which correspond to LDL and OxLDL, respectively, in gingival crevicular fluid. [ABSTRACT FROM AUTHOR]

Phage display technology is commonly applied for high‐throughput screening of single‐domain antibodies (sdAbs), and the problem of non‐specific adsorption caused by carrier proteins seriously affects the biopanning of single‐domain antibodies specific to haptens. In this paper, enrofloxacin (ENR)‐functionalized cryogels were prepared by the ethylenediamine (EDA) and carbodiimide methods for application in the biopanning of ENR‐specific phages. To improve the efficiency of biopanning, double blocking, a wash solution flow rate of 1 mL/min, and phage pre‐incubation were applied to the biopanning process through single‐factor experiments. Results of flat colony counting showed that the phage output of AG‐ENR cryogels was 15 times higher than that of AG cryogels for the same input amount. And seven complete sequences of ENR‐specific shark sdAbs were obtained by monoclonal phage ELISA and sequence alignment. All these results indicate that functionalized cryogels could be used as a novel and efficient method for phage biopanning for single‐domain antibodies to haptens. [ABSTRACT FROM AUTHOR]

This article focuses on the cigarette paper atrophy. This is an uncommon disease of unknown etiology. Recent studies have confirmed a significant increase of organ-specific autoantibodies in patients with lichen sclerosis et atrophiens (LSA). In the native African population LSA has been considered rare. Five patients are believed to be reported from West Africa. The disease is seen in children as well as in adults, and it occurs 10 times more frequently in women than in men, occuring around and after menopause. The anogenital region is involved in most cases. Nongenital lesions are symptomless and occur mostly on the trunk. The lesions start as small shiny ivory white papules that later coalesce to form well-demarcated plaques.

The article presents information on the evidences of the reactions resulting after the sciatic nerve ligation injury with respect to the correlation of CC-chemokine ligand 4 (CCL4) with the neuropathic pain behavior in the mice. It mentions the upregulation of the CCL4 mRNA at the nerve, and highlights the neutralization of the CCL4 with anti-CCL4 antibodies.

An increased level of granulocyte‐macrophage colony‐stimulating factor has a potential role in the development of autoimmune diseases, and the neutralization of its activity by monoclonal antibodies is a promising therapy for some diseases. Here, the crystal structure of the Fab region of EV1007, a fully human antibody expressed in Chinese hamster ovary cells that was developed from human peripheral blood mononuclear cells, is described. The structure closely resembles that of MB007, which is the Fab region of the same antibody expressed in Escherichia coli [Blech et al. (2012), Biochem. J.447, 205–215], except at the hinge regions between the immunoglobulin domains and the H3 loop region. This paper presents evidence for the flexibility of the hinge and H3 loop regions of the antibody based on the comparison of two independently solved crystal structures. [ABSTRACT FROM AUTHOR]

Rapid detection of antibodies to SARS‐CoV‐2 is critical for COVID‐19 diagnostics, epidemiological research, and studies related to vaccine evaluation. It is known that the nucleocapsid (N) is the most abundant protein of SARS‐CoV‐2 and can serve as an excellent biomarker due to its strong immunogenicity. This paper reports a rapid and ultrasensitive 3D biosensor for quantification of COVID‐19 antibodies in seconds via electrochemical transduction. This sensor consists of an array of three‐dimensional micro‐length‐scale electrode architecture that is fabricated by aerosol jet 3D printing, which is an additive manufacturing technique. The micropillar array is coated with N proteins via an intermediate layer of nano‐graphene and is integrated into a microfluidic channel to complete an electrochemical cell that uses antibody‐antigen interaction to detect the antibodies to the N protein. Due to the structural innovation in the electrode geometry, the sensing is achieved in seconds, and the sensor shows an excellent limit of detection of 13 fm and an optimal detection range of 100 fm to 1 nm. Furthermore, the sensor can be regenerated at least 10 times, which reduces the cost per test. This work provides a powerful platform for rapid screening of antibodies to SARS‐CoV‐2 after infection or vaccination. Highlights: This work demonstrates a microfluidic biosensor with 3D printed array electrodes that detects nucleocapsid (N) antibodies to SARS‐CoV‐2 in mere 10‐12 seconds. This breakthrough technology will save lives and help in the management of the ongoing COVID‐19 pandemic. [ABSTRACT FROM AUTHOR]

Introduction: Mast cells are regarded as a kind of classical anaphylaxis cells. However autoimmune diseases and allergic reactions have many similarities or overlaps. A large number of papers have proved that mast cells play a significant role in the pathogenesis of systemic lupus erythematosus (SLE). It is speculated that IgE, anti‐IgE antibodies, FcεRI, and anti‐FcεRI antibodies activate mast cells through autoimmune pathways and participate in the disease process of SLE. Naturally occurring protein molecules not only exist in monomer form, but also in polymer of protein molecules. Therefore, whether IgE, FcεRIα, anti‐IgE antibodies, and anti‐FcεRI antibodies also exist in polymeric forms in the natural state is worthy of further investigation. Methods: The serum samples and clinical data of 131 patients with SLE were collected from Qilu Hospital (Qingdao). Sixty healthy individuals were collected as the control group. Serum FcεRIα, anti‐IgE, and anti‐FcεRI were detected by enzyme‐linked immunosorbent assay. Serum IgE was detected by rate scatter nephelometry. A Chinese hamster ovarian cancer cell line CHO3D10 transfected with human FcεRIα was cultured and the cell protein extract was prepared. The existence forms of FcεRIα in the cell protein extract were detected by the native‐page method. Results: The serum FcεRIα in SLE patients was significantly higher than that in control group (3.52 [2.18, 4.71] µg/ml and 1.87 [1.52, 2.33] µg/ml, respectively; p <.05). Anti‐IgE was significantly lower than that in the control group (0.85 [0.55, 1.21] µg/ml and 1.23 [0.95, 1.58] µg/ml, respectively; p <.05). The CHO3D10 cell line expressed the FcεRIα, which had one kind of monomer (mFcεRIα) and two kinds of polymers (pFcεRIα) in the degeneration conditions. Conclusion: In patients with SLE, the expression of FcεRIα was increased and the level of anti‐IgE was decreased. FcεRIα had one kind of monomer and two kinds of polymers. Mast cell‐associated FcεRIα involved in the inflammatory lesion of SLE. [ABSTRACT FROM AUTHOR]

Selected current trends in analytical chemistry are discussed and traced back to original work published more than a hundred years ago in the same field. Gravimetric microanalysis has been transformed into a mass‐sensitive sensor technology. New developments in molecular spectroscopy are seen in Raman spectroscopy. In the area of chemical sensors, paper‐based devices with visual readout (dipsticks, microfluidic pads) celebrate a revival. In "green" applications, the separation of analytes from complicated matrices is often the key to successful analysis. Continuous separation techniques are essential for the 24/7 production of goods with certified quality. Finally, micro‐ and nanoscale techniques rely on highly selective receptors, and their development is based on detailed understanding of the structure of ligand–receptor complexes. Back to the future: Selected current developments in analytical chemistry are discussed in this Viewpoint and traced back to original work published up to 150 years ago. Discussed topics include selective mass determination, molecular spectroscopy, chemical sensors, separation methods, and bioinspired receptors such as antibodies. [ABSTRACT FROM AUTHOR]

OX40 is a costimulatory molecule that belongs to the tumor necrosis factor receptor (TNFR) superfamily. OX40 agonist‐based combinations are emerging as promising candidates for novel cancer immunotherapy. Clinical trials have shown that OX40 agonist antibodies could lead to better results in cancer patients. Using a hybridoma platform and three different types of immunization strategies, namely recombinant protein, DNA, and overexpressing cells, we identified a chimeric anti‐OX40 antibody (mAb035‐hIgG1 from DNA immunization) that shows excellent binding specificity, and slightly stronger activation of human memory CD4+ T cells and similar potent antitumor activity compared with BMS 986178, an anti‐OX40 antibody currently being evaluated for the treatment of solid tumors. This paper further systematically investigates the antigen‐specific immune response, the number of binders, epitope bins, and functional activities of antibodies among different immunization strategies. Interestingly, we found that different immunization strategies affect the biological activity of monoclonal antibodies. [ABSTRACT FROM AUTHOR]

A rapid method for collection of static crevicular fluid (SCF) on filter papers is described. Standardization of a Periotron® for volume quantitation of the SCF allowed accurate collection of crevicular fluid samples from periodontal disease patients. A method for elution of SCF was developed and was shown to provide nearly total recovery of antibody activity from the filter papers. Analysis of specific antibody to suspected periodontal pathogens in eight individuals indicated a site-to-site variation in antibody levels within the oral cavity. Also, SCF from certain sites exhibited antibody levels that were significantly greater than serum, supporting the concept of specific local antibody synthesis in gingival tissues. [ABSTRACT FROM AUTHOR]

New short-lived strains of mice (SAM-P), which have been developed by Takeda et al. (1981), show a defective antibody response to T dependent (TD) antigen in vitro, as demonstrated in the accompanying paper (see page 419). In the present study, we investigated the cellular site of the defect, using a cell culture system. In this paper, it is demonstrated that T-helper (Th) cell activity for the antibody response to TD antigen is impaired, while other cellular immune responses, e.g. mixed leucocyte reaction, cytotoxic T-lymphocyte response, and delayed-type hypersensitivity reaction, are normal. These results suggest that the defect in T-helper subset is limited in helper function for the antibody response, and that the helper function for the cell-mediated immune responses is intact. These two functions of the T-helper subset are apparently regulated in a different manner. The SAM-P strains of mice may thus serve as an appropriate model for studying functional heterogeneity in T-helper/inducer cell subsets. [ABSTRACT FROM AUTHOR]

An assay, the paper radioallergosorbent test (PRAST), for the measurement of specific serum IgE antibody in the ral is described in detail. This assay has been used, in conjunction with a modified PRAST for the determination of relative specific serum IgG antibody and the PRIST assay for total serum IgE [13], lo measure specific IgG and IgG and total IgE immune responses in normal parasite infected rats immunized using various protocols. The results indicate that there is a relationship between the basic IgE level and the immune response, i,e. a rat strain with a low constitutive IgE level demonstrates a weak response whereas a high level strain reacts strongly. When PRAST and passive cutaneous anaphylaxis (PCA) were compared, using standardized IgE antibody containing sera, the results were in good agreement. However, PRAST is the preferable assay as it shows less intrinsic variation, is more sensitive than PCA, and is not influenced by high serum IgE levels in the recipient animal. [ABSTRACT FROM AUTHOR]

The article presents a list of papers accepted for publication in forthcoming issues including "IgM Turnover in Chronic Liver Disease," by K. B. Jensen, S. Jarnum, L. Ranek and H. Westergaard, "Studies on the Induction of Antibody Synthesis Against Sulfanilic Acid in Rabbits," by B. Rubin, Macrophage Cytophilic Antibody to Diphtheria Toxoid in Desensitized Animals," by A. I. Kostiala and T. U. Kosunen and "The Identification of Immunoglobulin-Releasing Cells in Human Tonsillar Tissue," by K. M. Moriarty and D. S. Rowe.

Summary: Some human antibodies may paradoxically inhibit complement activation on bacteria and enhance pathogen survival in humans. This property was also claimed for IgG antibodies reacting with terminal galactose‐α‐1,3‐galactose (Galα3Gal; IgG anti‐αGal), a naturally occurring and abundant antibody in human plasma that targets numerous different pathogens. To reinvestigate these effects, we used IgG anti‐αGal affinity isolated from a pool of normal human IgG and human hypogammaglobulinaemia serum as a complement source. Flow cytometry was performed to examine antibody binding and complement deposition on pig erythrocytes, Escherichia coli O86 and Streptococcus pneumoniae serotype 9V. Specific nanobodies were used to block the effect of single complement factors and to delineate the complement pathways involved. IgG anti‐αGal was capable of activating the classical complement pathway on all the tested target cells. The degree of activation was exponentially related to the density of bound antibody on E. coli O86 and pig erythrocytes, but more linearly on S. pneumoniae 9V. The alternative pathway of complement amplified complement deposition. Deposited C3 fragments covered the activating IgG anti‐αGal, obstructing its detection and highlighting this as a likely general caveat in studies of antibody density and complement deposition. The inherent capacity for complement activation by the purified carbohydrate reactive IgG anti‐αGal was similar to that of normal human IgG. We propose that the previously reported complement inhibition by IgG anti‐αGal relates to suboptimal assay configurations, in contrast to the complement activating property of the antibodies demonstrated in this paper. [ABSTRACT FROM AUTHOR]

This case suggests a mechanistic rationale for the clinical efficacy of intravenous immunoglobulins (IVIG) in treating CD40 ligand (CD40L) deficiency associated neutropenia as it is the first reported instance of free and cell‐bound antineutrophil antibodies in a case of CD40L deficiency, accompanied by a prolonged and clinically severe neutropenia. [ABSTRACT FROM AUTHOR]

BACKGROUND: A short processing time and efficient scale‐up stem cell isolation bioprocess is essential to exploit the potential of these cells for the treatment of multiple chronic diseases. Various methodologies have been used for stem cell recovery. However, most of them present economic and/or time‐consuming drawbacks. RESULTS: This paper reports the characterization and optimization of immunoaffinity aqueous two‐phase systems (ATPS), a liquid‐liquid based separation technology enhanced with the PEGylation of the antibody as a promising platform in route to the separation of CD133+ stem cells. The best results were obtained with Ucon‐DEX or PEG‐DEX systems with tie‐line length (TLL) 15 or 20% w/w and volume ratio (VR) 3. When the PEGylated antibodies were added to these selected optimized ATPS, they partitioned to both phases, but with higher preference for the bottom phase. CONCLUSION: PEGylation was an effective strategy to induce a change in the partition behavior of the CD133/2‐biotin antibody when Ucon‐DEX ATPS were used. Similarly, variation of the parameters (TLL and VR) resulted to be useful for this goal. Nevertheless, further studies are required to find the optimal ATPS composition that will fractionate 100% of the antibody to the contaminants' opposite phase, making this system an ideal candidate to be tested for the selectivity of CD133+ stem cells. © 2019 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Monitoring of donor‐specific HLA antibodies (DSA) has become part of the clinical routine in kidney transplantation. This paper gives a brief overview on data from the Collaborative Transplant Study (CTS) and the Heidelberg Transplant Center on the clinical relevance of post‐transplant DSA monitoring in patients undergoing renal transplantation. The obtained findings underline the importance of DSA monitoring in the post‐operative course in immunologically high‐risk patients and patients with deterioration of graft function. Especially in patients with a pre‐activated immune system, a gap in the immunosuppressive therapy appear to lead to persistence, reappearance or de novo occurrence of strong, complement‐activating DSA, resulting in severe antibody‐mediated rejection (AMR) and, without timely intervention, in AMR‐related graft loss. [ABSTRACT FROM AUTHOR]

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