Column Agglutination Assay Using Polystyrene Microbeads for Rapid Detection of Antibodies against SARS-CoV-2.

Rapid serology platforms are essential in disease pandemics for a variety of applications, including epidemiological surveillance, contact tracing, vaccination monitoring, and primary diagnosis in resource-limited areas. Laboratory-based enzyme-linked immunosorbent assay (ELISA) platforms are inherently multistep processes that require trained personnel and are of relatively limited throughput. As an alternative, agglutination-based systems have been developed; however, they rely on donor red blood cells and are not yet available for high-throughput screening. Column agglutination tests are a mainstay of pretransfusion blood typing and can be performed at a range of scales, ranging from manual through to fully automated testing. Here, we describe a column agglutination test using colored microbeads coated with recombinant SARS-CoV-2 spike protein that agglutinates when incubated with serum samples collected from patients recently infected with SARS-CoV-2. After confirming specific agglutination, we optimized centrifugal force and time to distinguish samples from uninfected vs SARS-CoV-2-infected individuals and then showed concordant results against ELISA for 22 clinical samples, and also a set of serial bleeds from one donor at days 6-10 postinfection. Our study demonstrates the use of a simple, scalable, and rapid diagnostic platform that can be tailored to detect antibodies raised against SARS-CoV-2 and can be easily integrated with established laboratory frameworks worldwide.

Rapid Gel Card Agglutination Assays for Serological Analysis Following SARS-CoV-2 Infection in Humans.

High-throughput and rapid serology assays to detect the antibody response specific to severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) in human blood samples are urgently required to improve our understanding of the effects of COVID-19 across the world. Short-term applications include rapid case identification and contact tracing to limit viral spread, while population screening to determine the extent of viral infection across communities is a longer-term need. Assays developed to address these needs should match the ASSURED criteria. We have identified agglutination tests based on the commonly employed blood typing methods as a viable option. These blood typing tests are employed in hospitals worldwide, are high-throughput, fast (10-30 min), and automated in most cases. Herein, we describe the application of agglutination assays to SARS-CoV-2 serology testing by combining column agglutination testing with peptide-antibody bioconjugates, which facilitate red cell cross-linking only in the presence of plasma containing antibodies against SARS-CoV-2. This simple, rapid, and easily scalable approach has immediate application in SARS-CoV-2 serological testing and is a useful platform for assay development beyond the COVID-19 pandemic.

An approach to incompatible cross-matched red cells: Our experience in a major regional blood transfusion center at Kolkata, Eastern India.

INTRODUCTION: With the increased utilization of immunohematology (IH) analyzers in the transfusion medicine, type, and screen policy is the method of choice. Still, the importance of routine crossmatching could not be overruled. Here, we tried to understand the clinical conditions and safety of red cell transfusion and their outcomes. MATERIALS AND METHODS: This prospective study was conducted by IH laboratory, Medical College Kolkata, Blood Bank from October 1, 2015 to March 31, 2016. A set of 3cc ethylenediaminetetraacetic acid and clotted blood samples of the patients were received according to sample acceptance criteria. Blood grouping by conventional tube technique followed by crossmatching was performed by column agglutination technology (CAT) in polyspecific (IgG + C3d) gel media. Any positive result was rechecked in duplicate with additional two group-specific donor units. The persistent incompatibility was further evaluated using direct anti-human globulin test, auto control, antibody screening, and antibody identification by CAT. RESULTS: On the evaluation of 14,387 sets of patients' sample, only 100 were found to be incompatible (0.69%). Incompatibility rate is higher in females (59%). Eighty-five of these patients were repeatedly transfused. Only 38% of incompatible crossmatch were positive on indirect anti-human globulin test/antibody screening. Antibody could be identified in 16 of them. Seventeen of 100 incompatible samples (17%) presented with panagglutination, were managed with Rh, Kell phenotype/best-matched red cell units. In these 16 patients, 23 alloantibodies were identified; allo anti-E was the most common. CONCLUSION: This study showed antibody against the Rh system as the most common cause of incompatibility.

The Frequency Unexpected Antibodies at Kyungpook National University Hospital / 대한수혈학회지

BACKGROUND: Unexpected antibody screening and identification tests are very important pre-transfusion tests for preventing transfusion reactions. Nowadays, the column agglutination test is widely used in Korea. The results of many studies that used this method showed the decreased frequency of nonsignificant cold antibodies and an increased frequency of warm antibodies when compared with other studies that used the tube test or the microplate test. This study was performed in order to determine the accurate frequency and distribution of unexpected alloantibody by using the column agglutination test. METHODS: We analyzed the results from 32,218 antibody screening tests with using LISS/Coombs cards and ID-DiaCell I and II for the transfusion candidates and patients with hemolytic anemia who were seen at Kyungpook National University Hospital during a recent eight-year period. RESULTS: According to the results of the antibody screening test, 188 samples (0.58%) out of all 32,218 samples, were shown to be positive. Unexpected alloantibodies were detected in 86 patients (0.27%) with using the antibody identification test. The antibodies that were detected most frequently were anti-E (29 samples), followed by anti-D (8 samples), anti-M (8 samples) and anti-c (7 samples). CONCLUSION: The frequency and distribution of unexpected antibodies at our hospital are similar with those obtained in other Korean studies. The detection rates of warm antibodies, including Rh antibodies, were high. The proportion of Rh antibodies in patients with a gestation history was significantly higher than that in the patients without a gestation history. This study shows once again that pregnancy affects the antibodies and this supports the relationship between pregnancy and antibody formation.

Usefulness of column agglutination test for irregular antibody screening and identification / 대한임상병리학회지

BACKGROUND: Type and screen is recommended for efficient use of blood and reduction in workload in blood bank. Column agglutiation test is standardized and easy to perform, and provide clear and stable reactions that improve the interpretation of results. In this study, we compared column agglutination test(Ortho Diagnostic Systems Inc., USA) with conventional tube test and investigate its usefulness in irregular antibody screening and identification. METHODS: A total 182 samples were screened for irregular antibodies using column agglutination test and conventional tube test. And 18 patient's sera in which irregular antibodies previously screened by both tube and column agglutination tests were identified for irregular antibodies by tube and column agglutination tests. We compared the results of two tests. RESULTS: In the screening test, there was 96.7%(176/182) agreements between column agglutination test and conventional tube test. The column agglutination test showed stronger reactivity than tube test. In the irregular antibody identification, there was 88.8%(16/18) agreement between two tests and disagreement were seen in the identification of anti-P1 and anti-Leb antibodies. CONCLUSION: The results of column agglutination test are objective and superior to the conventional tube test in irregular antibody screening and identification tests. These results suggest that the column agglutination test will be useful and more convenient test in antibody screening and identification.

Discrepant Frequency of Rh Subtype and Kell Blood Group Antigens between Korean Pregnant Women and Their Neonates / 대한수혈학회지

BACKGROUND: The causes of hemolytic disease of the newborn are discrepancies of ABO group, Rh(D) or other RBC antigens. The discrepancies of Rh subgroups except Rh(D) and K typing can be seen rarely. The clinical symptoms of those types are mild and most clinicians have no interest in them. However, there exist some serious cases that need exchange transfusion. For that, we detected Rh subgroup phenotyping and Kell typing in blood obtained from cord and pregnant women and the frequency of discrepancy. METHODS: We examined the cord and mother's blood collected from 317 pregnant women from May to November, 1997. Rh(D) typing was done using slide method with anti-D (Dade, USA), and other Rh subgroup phenotyping using column agglutination test on MicroTyping system with Rh-K gel card (DiaMed, Switzerland). Irregular antibody screening was done in the cases of discrepancy. RESLUTS: The most frequent phenotype of Rh subgroup was CDe (41.3%) and then CcDEe (39.3%), but the K typing showed 0%. 90 cases of 317 pair-samples (28.4%) showed discrepancies between pregnant women and their neonates. The most frequent type of Rh discrepancy was c+E (50%) and then C or E (11.1%). 62 cord samples which obtained from neonates of Kyungpook and Chonnam provinces showed discrepancies, were all negative in the irregular antibody screening test. CONCLUSIONS: Rh subgroup phenotyping and irregular antibody screening in cord blood by column agglutination test is thought to be helpful in early diagnosis and treatment of hemolytic disease of the newborn, as a sufficient amount of cord blood can be collected easily rather than neonatal blood.

Irregular Antibody Screening in Cord Blood by Column Agglutination Test / 대한수혈학회지

BACKGROUND: Irregular antibodies are antibodies that are not regularly present in the serum of particular blood groups and its presence results in many problems including HDN (hemolytic disease of newborn) in transfusion medicine. Column agglutination test was recently introduced and has been widely used for advantages of standardized working procedures, standard reactions, stable reactions for hours and Coombs test without washing steps. We tested irregular antibodies in cord blood by column agglutination test and investigated its incidence and relation with HDN. METHODS: We tested the cord blood collected during delivery from 200 pregnant women. Column agglutination test was done on DiaMed ID MicroTyping System (DiaMed, Switzerland) and both LISS/Coombs and NaCl/Enzyme ID-cards were used. The antibody screening test was done first and antibody identification test was done to positive cases in same way. The cell typing and Rh phenotyping for cord blood of positive cases were also done. RESULTS: 2 cases of 200 samples (1%) were positive in the antibody screening test and each was identified as anti-D and anti-E antibody. CONCLUSIONS: Irregular antibody screening in cord blood by column agglutination test is thought to be helpful in early diagnosis and treatment of HDN.