Does decreasing the incubation period used in the antibody screen affect its sensitivity?

BACKGROUND: Pre-transfusion testing (PTT) encompasses a set of mandatory laboratory tests performed before red blood cell transfusion. The antibody screen, one component of PTT, commonly includes a 10-20 min incubation. The primary aim of this study was to determine if this period can be reduced when using current immunohematology methodologies. METHODS AND MATERIALS: Antibody screens were performed on reagent samples using Glass or Gel-based column agglutination technologies (CAT) and a solid phase red cell adherence (SPRCA) assay, with incubation periods of 1, 5, 10 and 15 min, and 20 min (SPRCA assay only). For each method, the shortest period producing a minimum of a 1+ reaction with all reagent samples was considered optimal. The sensitivity of each assay using the optimal period was calculated after performing antibody screens on 100 patient samples. RESULTS AND DISCUSSION: It was demonstrated that the incubation period in the SPRCA and Glass CAT systems can be reduced to 5 and 10 min, respectively, while achieving high assay sensitivity (98.9% in both). The incubation period in the Gel CAT system cannot be reduced from 15 min. Significant association between titre and reaction strength was observed for all three screening methods (p < 0.001 for both CAT methods, p = 0.041 for SPRCA). This study demonstrates that the incubation period used in the antibody screen can be reduced when using systems employing the Glass CAT and SPRCA methods, without affecting assay sensitivity. If confirmed, it could result in faster completion of PTT.

A prospective observational study to evaluate effect of heat inactivation on ABO titers performed by column agglutination technology and conventional tube technique.

BACKGROUND AND AIMS: When determining ABO antibody titers, immunoglobulin G (IgG) antibodies can be masked by immunoglobulin M (IgM) antibodies. Hence, the measurement of actual concentration of IgG requires methods like heat inactivation (HI) of plasma. This study was aimed at determining the effects of HI on IgM and IgG titers performed by conventional tube technique (CTT) and column agglutination technique (CAT). MATERIALS AND METHODS: This was a prospective, observational study conducted from October 2019 to March 2020. All consecutive A, B, and O group donors who gave consent for participation were included. All samples were consecutively tested by CTT and CAT, before and after HI (pCTT, pCAT). RESULTS: A total of 300 donors were included. IgG titers were found to be more than IgM titers. For group O, IgG titer results were higher for both anti-A and anti-B compared to group A and B. For group A, B, and O, pretreatment results were higher than posttreatment IgG titer results. Median anti-A titers were similar to median anti-B titers across all categories. Median IgM and IgG titers were higher for group O individuals than nongroup O individuals. There was reduction in IgG and IgM titers after HI of plasma. One log reduction in median titers was observed when ABO titers were performed by CAT and CTT. CONCLUSION: There is one log difference between median antibody titers estimated using heat inactivated and nonheat inactivated plasma. The use of HI for ABO isoagglutinin titer estimation can be considered in low resource settings.

Detection of ABO Discrepancy in a Case of Coronary Artery Disease by Conventional Tube Technique: A Miss by Column Agglutination Technology.

ABO and Rh blood grouping of donors and recipients is the first and foremost step in pretransfusion compatibility testing. Conventional tube technique (CTT) is used to test for blood grouping and Rh D typing. But the procedure is cumbersome, and there may be subjective variation during the interpretation of the test results. The other disadvantage is that it is not adaptable to automation. Many newer techniques, such as the column agglutination technique (CAT) used for pretransfusion testing, are amenable to automation. It is being preferred to shift from CTT to semiautomated or fully automated CAT platforms or other newer technologies in many blood centers. The CAT has the added advantage of increased sensitivity and stable end-point results. The results in automated platforms using CAT are equally efficient and reliable as CTT. However, sometimes it is noted that CAT misses subgroups detection. Here, we report a case with a subgroup of A that was failed to be detected by the CAT using dextran acrylamide gel, signifying the use of CTT in evaluating blood group discrepancy.

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.

Comparison between tube test and automated column agglutination technology on VISION Max for anti-A/B isoagglutinin titres: A multidimensional analysis.

BACKGROUND AND OBJECTIVES: VISION Max (Ortho Clinical Diagnostics, Raritan, NJ) measures anti-A/B isoagglutinin titres using automated column agglutination technology (CAT). We compared tube test (TT) and CAT of VISION Max comprehensively, including failure mode and effect analysis (FMEA), turnaround time (TAT) and cost, and suggested modified CAT (MCAT). MATERIALS AND METHODS: For 100 samples (each 25 for blood type A, B and O with anti-A and anti-B), anti-A/B isoagglutinin titres were measured by TT and CAT (1:2-1:1024 dilution), as well as by MCAT (with agglutination at 1:32 dilution, then perform additional testing from 1:64 to 1:1024). We assessed the agreement and correlation between TT and CAT and compared FMEA (risk priority number [RPN] score), TAT (h:min:sec) and cost (US dollar, US $) among TT, CAT and MCAT. RESULTS: TT and CAT showed overall substantial agreement (k = 0.73) and high correlation (ρ ≥ 0.75) except blood type O with anti-A (ρ = 0.68). Compared with TT, CAT showed lower RPN scores in FMEA and similar TAT and cost (FMEA, 33,700 vs. 184,300; TAT, 15:23:00 vs. 14:26:40; cost, 1377.4 vs. 1312.4, respectively). Regarding FMEA, TAT and cost, MCAT was superior to CAT or TT (43,810; 13:28:00; 899.2, respectively). CONCLUSION: This is the first multidimensional analysis on VISION Max CAT for measuring anti-A/B isoagglutinin titres. The results of anti-A/B isoagglutinin titres by CAT were comparable with those of TT. MCAT would be a safe, time-saving and cost-effective alternative to TT and CAT in high-volume blood bank laboratories.

COVID-19 Antibody Detection and Assay Performance Using Red Cell Agglutination.

Red cells can be labeled with peptides from the SARS-CoV-2 spike protein (C-19 kodecytes) and used as reagent cells for serologic screening of SARS-CoV-2 antibodies. We evaluated 140 convalescent COVID-19 donors and 275 healthy controls using C19-kodecytes. The analytical performance of the C19-kodecyte assay was compared with a virus neutralizing assay and two commercial chemiluminescent antibody tests (Total assay and IgG assay, Ortho). The C19-kodecyte assay detected SARS-CoV-2 antibodies with a sensitivity of 92.8% and specificity of 96.3%, well within the minimum performance range required by FDA for EUA authorization of serologic tests. The Cohen's kappa coefficient was 0.90 indicating an almost perfect agreement with the Total assay. The Spearman's correlation coefficient was 0.20 with the neutralizing assay (0.49 with IgG, and 0.41 with Total assays). The limited correlation in assay reaction strengths suggested that the assays may be influenced by different antibody specificities. The C19-kodecyte assay is easily scalable and may vastly improve test capacity in any blood typing laboratory using its routine column agglutination platforms. IMPORTANCE We recently developed a red cell based assay to detect SARS-CoV-2 antibodies in human plasma. In the current study, we show the hands-on application of this assay in a group of COVID-19 convalescent plasma donors and healthy individuals. We compared our assay against three published assays, including two that are widely used for patient care in the United States. Our assay compared well with all three assays. Our easily scalable assay can be used for population-wide screening of SARS-CoV-2 antibody status. It can be readily established in any hospital blood bank worldwide using its routine equipment.

High-sensitivity detection of clinically significant red blood cell antibodies by the column agglutination technique.

BACKGROUND: The detection of clinically significant antibodies to red blood cell antigens is important for the selection of compatible blood for patients. The conventional test tube technique (CTT) is commonly used as the gold standard test, but manual testing and visual detection of hemagglutination may produce errors. A more recently developed method, the column agglutination technique (CAT), facilitates ease of testing. OBJECTIVES: To investigate the specificity and sensitivity of the CAT compared to the CTT for the screening of clinically significant antibodies from the Rh blood group. MATERIAL AND METHODS: Standard antibodies to the Rh blood group, anti-D, -C and -E, were used as examples of clinically significant antibodies in transfusion science. The antibodies were serially diluted by two-fold, then reacted with screening cells with different antigen expression. The hemagglutination reaction was investigated using both techniques, and the grades and scores of the reactions were used to analyze the sensitivity and specificity of the CTT and CAT. RESULTS: The CAT had a better sensitivity than the CTT. The lowest antibody dilution of 1:8192 could be detected using CAT, while a dilution of only 1:2048 could be detected with CTT. However, the CTT and CAT were equal in specificity. The 2 techniques specifically detected all antibodies to the screening cells. CONCLUSIONS: Both the CAT and CTT showed 100% specificity. However, the CAT exhibited more sensitivity than the CTT, and can be used in substitution of, or in parallel with, the CTT technique for red blood cell phenotyping, antibody screening, identification, and crossmatching.

Prognostic Significance and Prevalence of IgG Subtypes in Rh Haemolytic Disease of Fetus and Newborn.

Many authors have reported poor prognostic value of anti-D antibody titer in the setting of Hemolytic Disease of Fetus and Newborn (HDFN). According to literature, HDFN cases with IgG1 and IgG3 have more severity compared to IgG2 and IgG4.Therefore, we planned this study to evaluate the prevalence and prognostic value of IgG subtypes in the setting of Rh HDFN. This was a retrospective study performed at a tertiary care center in north India from October 2015 to November 2017. Women with anti-D antibody were included in the study and categorized on the basis of presence of specific IgG subtype. "DAT IgG1/IgG3 ID" card (BIO-RAD) was used for determining the subclass of IgG. Various clinical, laboratory & interventional parameters were used to categorize fetal outcome in severe and non-severe cases. Perinatal outcome was then compared between women with different IgG subclass profile. Subclass distribution among 80 alloimmunized women was 26.2% for IgG1, 15% for IgG3, 46.2% for IgG1 + IgG3 and the rest had neither IgG1 nor IgG3. Severity of HDFN was significantly higher when IgG1 &/or IgG3 were present alone or in combination, compared to cases with absence of IgG1 or IgG3 (p value < 0.05). Risk of severe HDFN was significantly higher in the presence of IgG1 &/or IgG3 and the severity was highest when both IgG1 and IgG3 were present. We recommend that IgG subclass determination should be included in a multi-parameter protocol for more accurate prediction HDFN severity to ensure timely referral and intervention.

Comparative evaluation of DTT treated ABO isoagglutinin titres performed by two methods with solid phase red cell adhesion (SPRCA) titres.

BACKGROUND AND AIMS: Measurement of actual concentration of IgG requires methods like treatment of serum with dithiothreitol (DTT). This study was aimed at comparing of DTT treated ABO titres performed by conventional test tube technique (CTT) and column agglutination technique (CAT) with HA/SPRCA. MATERIALS AND METHODS: This was a prospective, observational study conducted from October 2019 to March 2020. All consecutive A, B and O group donors who gave consent for participation were included. All samples were tested by CTT and CAT before and after DTT treatment (pCTT, pCAT) and with HA/SPRCA. RESULTS: A total of 300 donors were included; 100 each from A, B and O blood group donors. Group O titres were higher than group A/B titres. Group O titres were highest when performed by pCAT, followed by pCTT and lowest by HA/SPRCA. Group A/B titres were highest when performed by HA/SPRCA, followed by pCAT and pCTT for anti-A and highest when performed by pCAT, followed by HA/SPRCA and lowest by pCTT for anti-B. CONCLUSION: Results obtained by pCAT were closer to results obtained by pCTT, whereas those obtained by HA/SPRCA were variable. SPRCA offers the advantage of automation, no inter-observer variation and less time consumption because IgM interference is not observed with SPRCA, thus providing an alternative to pCTT. However, these methods cannot be used interchangeably and to discern the most suitable method, a clinical impact of these results needs to be studied.

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.

A comparison of three column agglutination tests for red blood cell alloantibody identification.

OBJECTIVE: Commercial kits of column tests for pre-transfusion testing have progressively replaced conventional tube tests in most laboratories. Aim of this study was to compare three commercial test cell panels for the identification of irregular red blood cell (RBC) alloantibodies. Overall, 44 samples with a positive indirect antiglobulin test (IAT) by routine testing were used for comparison of following panels: Ortho RESOLVE® panelC (Ortho Clinical Diagnostics (OCD), Milan, Italy), ID-DiaPanel(-P) (Bio-Rad Laboratories, CA, USA) and Identisera Diana(P) (Grifols, Barcelona, Spain). Column agglutination techniques were used, with microtubes containing either microgel (Bio-Rad/Grifols) or glass bead microparticles (Ortho). RESULTS: Alloantibody identification was possible in 38 samples, of which identical identification was shown in 33 samples by all methods. The remaining samples showed differences between certain methods, with the gel card system being superior to the glass card system for analyzing stored samples Considering that not all samples were evaluated in all three methods, the concordance rate reached 100% between Bio-Rad and Grifols, 90.5% between Bio-Rad and OCD, 86.5% between OCD and Grifols and 90.5% between all methods. Although differences in sensitivities were seen for specific antibodies, the three methods showed comparable performance for the identification of RBC alloantibodies.

Defining critical antibody titre in column agglutination method to guide fetal surveillance.

INTRODUCTION: A critical anti D antibody titre, defined for the conventional tube method of Indirect Coomb's test (ICT), when employed in the more sensitive column method could result in unnecessary referrals and frequent obstetric doppler scans. This study aimed to compare anti D titres by tube and column method in antenatal mothers, to assess their correlation with fetal anemia and to determine a critical titre for the column method. METHODS: Forty six antenatal mothers with anti D antibody were included in the study. Antibody titration was performed by serial twofold dilution of serum by both column and tube method and were correlated with middle cerebral artery peak systolic velocity (MCA PSV) measurement by Doppler ultrasonography. Receiver operating curve (ROC) was used to determine the cut-offs for critical titre by tube and column method in predicting fetal anemia. RESULTS: Column method had a median titre 3 fold higher than tube method. There was a significant association between fetal anemia by USG with median critical titres determined for both column (p = 0.031) and tube method (p = 0.016). ROC analysis showed the cut off for critical titres in column method as 64 with 90 % sensitivity, 72.7 % specificity and 75.38 % accuracy. CONCLUSIONS: The use of critical titre for anti D antibody, defined for the tube method, when applied to the column agglutination method would lead to increased referrals to specialized fetal medicine centres. Rather, an Anti D titre of 64 by column method can predict the likelihood of fetal anemia and should be considered as the critical titre to guide patient referrals.

Distribution of antenatal alloimmunization in the southern districts of West Bengal and its significant associated factor.

OBJECTIVES: Detection of maternal irregular antibodies against red blood cell antigen is vital in the management of hemolytic disease of fetus and newborn. There are no uniform guidelines related to antenatal antibody screening and identification in the developing Country like India. This study was aimed to identify such alloimmunization and its associations. MATERIALS AND METHODS: This prospective study was conducted on antenatal mothers at a tertiary care center. The mothers having a history of anti-D administration, blood transfusion, and autoimmune disorders were excluded from the study. Initial indirect antiglobulin test (IAT) was performed in all blood samples by conventional tube technique (CTT) to identify alloimmunization. IAT-positive samples were screened for irregular antibody by column agglutination technology (CAT). Antibody screen-positive samples were further analyzed in 11-cell panel by CAT. Antibody strength was measured by serial double dilution by CTT. The source of isoimmunization was identified by extended Rh phenotype of women, husband, and newborn. RESULTS: A total of 12 (2.3%) women out of 530 were positive for IAT and antibody screen. Antibody could be identified in 11 women, of which anti-D (5) was the most common, followed by anti-C + anti-D (4), anti-C + anti-E (1), and anti-C (1). All four cases of anti-D + anti-C were distinguished from anti-G by differential adsorption and elution. There was a significant association with alloimmunization versus increased gravid status, antepartum hemorrhage, and past history of newborns with neonatal jaundice. CONCLUSION: All pregnant women with history of antepartum haemorrhage, newborn with neonatal jundice should be screened for alloantibody for early detection and better management of HDFN.

Comparison of a column agglutination technology-based automated immunohematology analyzer and a semiautomated system in pretransfusion testing.

INTRODUCTION: Semi-automated equipment using Column agglutination technology (CAT) is widely used where centrifugation and incubation are automated but substantial amount of the work is still executed manually. Larger laboratories are moving towards automation to eliminate errors, reducing exposure to bio-hazardous samples, assuring traceability, reliability, turnaround time (TAT) and throughput. Moving towards automation and greater reliability, we therefore, decided to install an automated immunohematological (IH) analyzer "Vision". In this study we evaluated reliability and performance before clearing "Vision" for routine use. MATERIALS AND METHODS: Study was conducted in the Department of Transfusion-Medicine. The primary objective was to assess the reliability of results and compare with routine use semi-automated BIOVUE system (Reference system). Secondary objective was to evaluate the performance (TAT and throughput) of the Vision to handle routine and emergency workload. RESULTS: Total of 1276 known samples were used to assess 2640 pre-transfusion tests (1229 ABO/Rh D typing; 1229 antibody screening; 54 antibody identification; 86 crossmatch and 42 DAT). All 1229 ABO Rh typing results were concordant between the two systems. Overall agreement between the Vision IH analyzer and reference system for ABO Rh typing was 99.95%. All antibody screening, crossmatch and DAT results were concordant between the two systems. TAT of Vision was substantially shorter than the reference system for all test profiles. CONCLUSION: Based on study results, Vision was approved for routine use in laboratory. It was found to be reliable with considerably shorter TAT and capable of handling high throughput of immunohematological tests.

Comparison of the tube test and column agglutination techniques for anti-A/-B antibody titration in healthy individuals.

BACKGROUND AND OBJECTIVES: Determination of the anti-A/-B titre pre- and post-transplantation is beneficial for treatment selection. Currently, the recommended method for antibody titration is the tube test (TT) assay. Dithiothreitol (DTT) is used for IgM antibody inactivation. Recently, a fully automated antibody titration assay using the column agglutination technique (CAT) was developed (auto-CAT). Our aim was to compare the auto-CAT and TT techniques for ABO antibody titration, to evaluate the effectiveness of DTT-treated plasma for use with auto-CAT and to define the cut-off value for antibody titration by auto-CAT. MATERIALS AND METHODS: We enrolled 30 healthy individuals, including 10 each for blood types A, B and O. We performed antibody titre measurement using the TT technique and auto-CAT simultaneously. Auto-CAT uses the bead column agglutination technology. RESULTS: With the auto-CAT cut-off value set to weak (w)+ with DTT treatment plasma, the concordance rate was 45%, and the weighted kappa value between TT and auto-CAT results was 0·994 in all subjects. Furthermore, there was a significant positive correlation between the anti-A/-B titre results obtained using the TT technique and auto-CAT in all blood types. Moreover, a positive bias (falsely elevated end-points due to agglomeration of A/B cells) was not observed in auto-CAT testing using DTT-treated plasma. CONCLUSION: Our results show that 1+ agglutination using the TT technique is equivalent to w+ agglutination obtained using auto-CAT. We recommend that DTT may be used with auto-CAT to measure antibody titres. Thus, we suggest that auto-CAT is useful for antibody titration in routine examination.

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.

Evaluation of new indigenous "point-of-care" ABO and Rh grouping device.

BACKGROUND: Erycard 2.0 is a "point-of-care" device that is primarily being used for patient blood grouping before transfusion. MATERIALS AND METHODS: Erycard 2.0 was compared with conventional slide technology for accuracy and time taken for ABO and Rh forward grouping result with column agglutination technology (CAT) being the gold standard. Erycard 2.0 as a device was also evaluated for its stability under different storage conditions and stability of result till 48 h. In addition, grouping of hemolyzed samples was also tested with Erycard 2.0. Ease of use of Erycard 2.0 was evaluated with a survey among paramedical staff. RESULTS: Erycard 2.0 demonstrated 100% concordance with CAT as compared with slide technique (98.9%). Mean time taken per test by Erycard 2.0 and slide technique was 5.13 min and 1.7 min, respectively. After pretesting storage under different temperature and humidity conditions, Erycard 2.0 did not show any deviation from the result. The result did not change even after 48 h of testing and storage under room temperature. 100% concordance was recorded between pre- and post-hemolyzed blood grouping. Ease of use survey revealed that Erycard 2.0 was more acceptable to paramedical staff for its simplicity, objectivity, and performance than conventional slide technique. CONCLUSION: Erycard 2.0 can be used as "point-of-care" device for blood donor screening for ABO and Rh blood group and can possibly replace conventional slide technique.

Comparison of antibody titers using conventional tube technique versus column agglutination technique in ABO blood group incompatible renal transplant.

INTRODUCTION: Measurement of alloantibody titer to a red cell antigen (ABO titers) is an integral part of management of ABO incompatible kidney transplants (ABOiKT). MATERIAL AND METHODS: There are different methods of titer estimation. Alloantibody detection by tube titration and Gel agglutination columns are accepted methodologies. It is essential to find the difference in titers between the two methods so as to set the 'cut-off' titer accordingly, depending upon the method used. RESULTS: We did a prospective observational study to compare and correlate the ABO titers using these two different techniques - conventional tube technique (CTT) and the newer column agglutination technique (CAT). A total of 67 samples were processed in parallel for anti-A/B antibodies by both tube dilution and column agglutination methods. The mean titer by conventional tube method was 38.5 + 96.6 and by the column agglutination test was 96.4 + 225. The samples correlated well with Spearman rho correlation coefficient of 0.94 (P = 0.01). CONCLUSION: The column agglutination method for anti A/B titer estimation in an ABO incompatible kidney transplant is more sensitive, with the column agglutination results being approximately two and half fold higher (one more dilution) than that of tube method.

Can serological methods help distinguish between prophylactic and alloimmune anti-D?

OBJECTIVES: Enzyme indirect antiglobulin test (EIAT) and polyethylene glycol IAT (PIAT) were evaluated for their potential use as tests to distinguish between prophylactic and alloimmune anti-D in plasma by comparing with a tube variation of the standard low ionic strength solution-IAT (LISS-IAT). BACKGROUND: Laboratories performing the screening of RhD-negative pregnant women are required to provide clinicians with guidance as to the source of detected RhD antibodies. Currently, this is derived from RhIg immunoprophylaxis history, agglutination scores and titration results, where performed. A serological test that can differentiate between prophylactic and alloimmune anti-D would be useful in the diagnosis of RhD alloimmunisation in pregnant women. MATERIALS AND METHODS: Plasma samples (n = 273) [fresh (collected from April 2014 to February 2015) and frozen (up to 2 years)] from antenatal females, preoperative males and females over child-bearing age were used in this study. Samples were identified as containing anti-D by routine column agglutination (CAT) and were tested by tube LISS-IAT, EIAT and PIAT, and a score difference was calculated. RESULTS: A total of 32% of alloimmune anti-D samples demonstrated an increase in agglutination score (+2 or +3) when tested by EIAT. A significant increase in agglutination score for alloimmune samples using EIAT compared with LISS-IAT was observed. EIAT had a sensitivity (Sn) of 59%, positive predictive value (PPV) of 100% and specificity (Sp) of 100% for alloimmune anti-D. CONCLUSION: EIAT is capable of confirming but not excluding the presence of alloimmune anti-D in samples where anti-D is detected in routine antibody screening.

Evaluation of the Automated Immunohematology Analyzer ORTHO VISION for ABO Antibody Titration / 대한수혈학회지

BACKGROUND: ABO antibody titration is important in cases such as ABO incompatible hemolytic disease of the fetus and newborn (HDFN), ABO incompatible bone marrow, or solid organ transplantation. This study was conducted in order to evaluate usability of ORTHO VISION (Ortho Clinical Diagnostics, Raritan, USA) designed automated ABO antibody titration equipment. METHODS: The isoagglutination titers were determined in 80 subjects (20 A, 20 B, 40 O (anti-A 20, anti-B 20)) using a conventional tube technique, including a 30 minute room temperature phase (CTT), Dithiothreitol treated manual column agglutination technique (MCAT), and automated column agglutination technique (ACAT) by ORTHO VISION. The concordance of titer was compared within one dilution step between the two methods. RESULTS: The isoagglutinin titers measured by the ACAT with anti-human globulin poly cassette (ACAT_Poly) and anti-human globulin IgG cassette (ACAT_IgG) were the highest and the isoagglutinin titer measured by the MCAT was also higher than that by the CTT. The isoagglutinin titer measured by the ACAT with reverse diluents cassette (ACAT_Reverse) was similar to that measured by the CTT. The concordance of anti-A and anti-B titers between CTT and ACAT_Reverse was 83% and 68%. The concordance of anti-A and anti-B titers between MCAT and ACAT_Poly was 100% and 83%. The concordance of anti-A and anti-B titers between MCAT and ACAT_IgG was 98% and 88%. CONCLUSION: Automated isoagglutinin titration using ACAT_Poly or ACAT_IgG without DTT showed reliable concordance with DTT treated MCAT, and it appears to be a possible replacement for the conventional MCAT method.