Antigen-positive Blood Research Articles

Serological analysis of autoantibody mimicking anti-Rh(e) specificity: a case report

Mimicking antibodies are described as autoantibodies with the “wrong” specificity, possessing both specificity and panreactivity. We report the case of a female patient with suspected hemolytic anemia in whom an autoantibody mimicking anti-Rh(e) specificity was found in both plasma and red blood cell samples. Mimicking antibodies are not usually identified in the clinic; however, it is still necessary to determine the specificity of mimicking antibodies to avoid antigen-positive blood transfusions and achieve maximum transfusion compatibility.

Hepatitis delta virus infection in Turkey: A meta-analysis of prevalence

ObjectivesHepatitis delta virus (HDV) infection has been granted orphan disease status by the US Food and Drug Administration and the European Medicines Agency owing to its rarity and relatively limited research and treatment options. Turkey is considered an endemic country for the virus. We aimed to provide a current and updated country- and region-specific HDV infection prevalence. MethodsIn this meta-analysis, we searched databases, including MEDLINE, PUBMED, EMBASE, and UlakBim (Turkish Medical Index) published between January 1, 2006, and December 31, 2022. We included blood donor studies, outpatient clinic studies that comprised patients without cirrhosis, and inpatient clinical studies that comprised patients with cirrhosis. Turkey was divided into three regions: West, Central, and East Turkey. ResultsAfter a systematic assessment, 41 studies were included. Using a random-effects model, the estimated HDV prevalence among hepatitis B surface antigen–positive blood donors, outpatient clinic, and inpatient clinic patients were 3.37% (confidence interval [CI] 1.99-6.11), 5.05% (CI 4.00-6.23), and 29.06% (CI 10.45-51.79), respectively. The HDV prevalence among outpatient clinic patients in Western, Central, and Eastern regions were 3.38% (CI 2.47-4.44), 2.15% (CI 1.37-3.09), and 9.81% (CI 6.61-13.55), respectively. ConclusionsEast Turkey continues to have a high burden of HDV. Public health efforts, such as screening, should be targeted accordingly.

Genetic Characterization of the Dengue Virus Type 3 Genotype I Prevailing in Dhaka, Bangladesh, 2021.

Background: In Bangladesh, dengue has been prevalent since its resurgence in 2018, and the dominant causative virus in 2019 was considered dengue virus serotype 3 (DENV-3). However, limited information is available for DENV serotype/genotype circulating after 2020. Materials and Methods: Viral RNA was extracted from NS1 antigen-positive blood samples of febrile patients in Dhaka, in 2021. DENV gene was detected by semi-nested RT-PCR, and sequences of envelope (E) gene and C-prM gene were determined by direct sequencing of RT-PCR products for genetic analysis. Results: Among 172 NS1-positive samples collected, 91 samples were assigned to DENV-3 and DENV-2 (88 and 3 samples, respectively) by RT-PCR targeting the C-prM gene. Phylogenetic analysis of the E gene for the 17 representative DENV-3 samples showed that all the viruses belonged to genotype I, forming a cluster (B-cluster) with those of DENV-3 reported in Bangladesh in 2017. Analysis of the deduced amino acid sequences of E protein revealed 16 amino acid substitutions, including two novel ones (G221W, L285P), and a substitution T223I that was specifically found in DENV-3 B-cluster. Conclusion: This study showed the persistent predominance of DENV-3 genotype I in Bangladesh having unique genetic traits in the E gene. (Approval number: MMC/IRB/2022/468).

RHESUS (RH) AND KELL ANTIGEN INCOMPATIBILITY – INDICATIONS FOR PREIMPLANTATION GENETIC TESTING

Rh or Kell incompatibility occurs when an antigen-negative woman is exposed to antigen-positive blood cells during pregnancy or transfusion. After the initial exposure, antibodies persist and cross the placenta into fetal circulation in subsequent pregnancies, causing destruction of antigen-positive fetal red blood cells1, or suppressing the production of fetal red blood cells.2 Complications can include miscarriage, stillbirth, fetal or neonatal anemia, or hyperbilirubinemia.1,2 If the male partner is heterozygous, preimplantation genetic testing (PGT) for monogenic conditions (PGT-M) is an option to identify embryos that are Rh or Kell-negative, preventing incompatibility and associated complications.

A retrospective study on the prevalence of weak D antigen (Du) in a blood bank in a tertiary care hospital in Maharashtra, India

Background: One of the most immunogenic blood groups known to humans is the Rhesus (Rh) blood group. The Rh blood group with variable expression of D antigen is one of the complex systems in immunohematology. Sometimes, the variable expression of RhD antigen leads to the presence of weaker forms that are not detected by routine laboratory procedures. Aims: The study was conducted to determine the frequency of RhD negativity and weak D antigens. Settings and Design: This retrospective study was conducted in the Department of Immunohematology and Transfusion Medicine at a tertiary care center hospital for three years from June 2017 to June 2020. Materials and Methods: All the serial Rh grouping tests were conducted by the gel technique or by the tube method with two Anti-D reagents. Negative D antigen samples were further tested for weak D antigen by Coombs Test. Statistical analysis used the Statistical Package for Social Sciences (SPSS). Results: Out of a total of 17262 samples, 15400 (89.2%) tested Rh-positive, and 1866 (10.8%) tested Rh-negative. Weak D was positive in 52(0.027%) samples out of the RhD negative samples. Conclusion: The frequency of weak D antigen is low, but the strong immunogenicity of RhD antigen calls for the need for appropriate testing. This is of concern in RhD negative pregnant females, as it can cause alloimmunization if accidentally weak D antigen-positive blood is transfused.

Testing for weak D Antigen: Spectrum and its applied role in rhesus-negative transfusions in Andaman and Nicobar Islands.

Objectives:Rhesus (Rh) blood group with variable expression of D antigen is one of the complex systems in immunohematology. Weak D antigen is a phenotype where the D antigen is weakly expressed on red blood cells, and this antigen cannot be detected by routine methods. This study was conducted to determine the frequency of Rh D negativity and weak D antigen among healthy blood donors and to review the clinical significance of weak D antigen pertaining to Rh D-negative transfusions.Materials and Methods:This cross-sectional prospective study was conducted in G. B Pant Hospital from January 2016 to June 2017 in which all the blood donors from Port Blair and adjacent islands of Andaman and Nicobar were grouped for Rh D antigen and those who tested negative for the D antigen were further tested for weak D antigen by incubating for 30 min and subsequent addition of anti-human globulin sera.Results:Out of 6415 donors, 6085 (94.86%) were Rh D positive and 330 (05.14%) were Rh D negative. Among the Rh D-negative donors, 05 (01.51%) were positive for weak D antigen. The frequency of Rh D negativity was 25.76% in a blood group, 25.15% in B, 07.88% in AB and 41.21% in O blood group phenotype.Conclusion:Although the frequency of weak D antigen is low (01.51%), the strong immunogenicity of Rh D antigen discernates the need for appropriate testing for weak D antigen. This is of particular concern in Rh D-negative pregnant females as it can produce alloimmunization if accidentally given weak D antigen positive blood.

Hepatitis B Virus Serum DNA andRNA Levels in Nucleos(t)ide Analog-Treated or Untreated Patients During Chronic and Acute Infection.

Treatment of chronic hepatitis B (CHB) patients with nucleos(t)ide analogs (NAs) suppresses hepatitis B virus (HBV) DNA synthesis but does not affect synthesis of HBV pregenomic RNA (pgRNA). Hepatitis B virus pgRNA is detectable in the serum during NA treatment and has been proposed as a marker of HBV covalently closed circular DNA activity within the infected hepatocyte. We developed an automated assay for the quantification of serum HBV pgRNA using a dual-target real-time quantitative PCR approach on the Abbott m2000sp/rt system. We demonstrate accurate detection and quantification of serum HBV RNA. Hepatitis B virus DNA was quantified using the Abbott RealTime HBV viral load assay. We further compared serum nucleic acid levels and kinetics in HBV-positive populations. Samples included on-therapy CHB samples (n = 16), samples (n = 89) from 10 treatment naïve CHB subjects receiving 12 weeks of NA treatment with 8-week follow-up, hepatitis B surface antigen-positive blood donor samples (n = 102), and three seroconversion series from plasmapheresis donors (n = 79 samples). Conclusion: During NA treatment of CHB subjects, we observed low correlation of HBV DNA to pgRNA levels; pgRNA concentration was generally higher than HBV DNA concentrations. In contrast, when NA treatment was absent we observed serum pgRNA at concentrations that correlated to HBV DNA and were approximately 2 log lower than HBV DNA. Importantly, we observe this trend in untreated subject samples from both chronic infections and throughout seroconversion during acute infection. Results demonstrate that the presence of pgRNA in serum is part of the HBV lifecycle; constant relative detection of pgRNA and HBV DNA in the serum is suggestive of a linked mechanism for egress for HBV DNA or pgRNA containing virions.

Guideline on the management of acute chest syndrome in sickle cell disease.

1. Department of Haematology, Guy’s and St Thomas’ NHS Foundation Trust 2. Lane Fox Respiratory Unit, St Thomas’ Hospital, Guy’s and St Thomas’ NHS Foundation Trust 3. Department of Haematology, North Middlesex Hospital 4. Department of Paediatric Haematology, Bristol Royal Hospital for Children 5. Department of Haematological Medicine, King’s College Hospital 6. Department of Haematology, Whittington Hospital

Bluetongue virus serotype-1 in goats in the Pithoragarh area of Uttarakahand, India.

This short communication reports the results of a bluetongue sero-surveillance conducted in the Pithoragarh hills of Uttarakhand in India during the autumn of 2011. Unclotted blood and serum samples were collected from 51 goats for detection of bluetongue virus (BTV) antigen and antibodies. Of the 51 collected samples, 18 (35%) were positive to an indirect ELISA and 33 (64%) resulted positive to a BTV ELISA antigen. From a strong antigen-positive blood sample, a BTV was isolated (named as PTG-13) on cell culture and was subsequently confirmed as BTV-1 by RT-PCR and partial sequencing of genome segment-2. The goat serum samples were found to contain high titer of neutralising antibodies against BTV-23, nonetheless the virus could not be isolated. Interestingly, no neutralizing antibodies were detected against PTG-13 or other BTV-1 isolate, which suggests that sampling was probably done before the development of neutralizing antibodies against PTG-13 virus in the host. Isolation of BTV-1 (PTG-13) and presence of BTV-23 neutralizing antibodies in serum samples indicate that goats were probably infected with BTV-1 and 23 in different periods.

Cytomegalovirus-specific antibodies to an immediate early antigen and a late membrane antigen and their possible role in controlling secondary cytomegalovirus infection

In 24 renal transplant recipients who had a secondary cytomegalovirus (CMV) infection, the magnitude and development of CMV-specific antibodies directed against two different antigens were studied in relation to the presence of CMV-immediate early antigen-positive peripheral blood leucocytes (CMV antigenaemia). These antibodies were measured in an antigen-capture ELISA using two monoclonal antibodies: one directed against the major immediate early antigen (IEA) and a second one directed against the CMV-encoded glycoprotein B (gB). A statistically significant inverse relationship between the level of anti-IEA antibodies present at the time of transplantation as well as the magnitude of the increase of these antibodies during CMV infection and the maximum number of IEA-positive cells during infection was shown. In contrast, both anti-gB and anti-total CMV antibodies did not give any correlation with the CMV antigenaemia. This may indicate that the anti-IEA immune response plays a role in defence mechanisms against a CMV infection.

一般病院における緊急輸血体制の確立にむけて〜特にＯ型緊急輸血について〜

To prevent delays in transfusion and the development of hemolytic transfusion reactions at the emergency room of a local hospital, we implemented a Type O blood usage program for life-threatening bleeding patients in August 2001 according to Japanese government guidelines for transfusion. To better understand actual conditions for urgent blood transfusion before and after implementation of this program, we surveyed the time between ordering to the start of transfusion, shock index at the start of transfusion, total volume of transfused blood, causes of death, irradiation of blood, side effects, and reasonable adaptation of transfusion. Results showed the time between ordering to the start of transfusion shortened to 9.3 minutes from 62.2 minutes after implementation. There was no case in which non-irradiated blood unit, was transfused, nor of incompatible blood transfusion following cross matching. Further, there was no increase in workload following implementation of the Type O blood usage program. In conclusion, Type O blood usage programs can dramatically save time between ordering to the start of transfusion. However, they carry risks such as Rh-positive blood transfusion to Rh-negative patients (about 0.5%) and antigen-positive blood transfusion to patients positive for irregular antibodies (about 1.3%). Adaptation of this transfusion must be carefully controlled. All patients receiving type O blood transfusion should be investigated for suitability after transfusion.

交差適合試験を実施しない時間外緊急輸血マニュアル

We introduced an automated instrument for pretransfusion testing during off-hours based on column agglutination technology. As the time required for testing ABO group and Rh type is at least 20 minutes, we also prepared manuals regarding emergency transfusion of uncrossmatched blood. During a two-year period, 25 patients received 209 units (8.4 units per patient) of group O blood, and 19 patients received 289 units (15.2 units per patient) of group-specific blood. Additionally, 5 patients received 98 units of group-specific blood following group O blood. Fourteen patients (26.9%) required massive transfusion (20 or more units of blood within 24 hours). Accurate blood typing was carried out by using a blood sample drawn from the patient prior to infusion of group O blood. Antibody screening was carried out using this specimen the next day, and if a patient was positive for an antibody, complete compatibility testing was also performed. All patients were followed clinically and for any signs of transfusion reactions or crossmatching difficulties, but none developed. Although one patient had anti-E antibody present on admission and received E antigen-positive blood, no elevation of antibody level or delayed hemolytic reaction occurred. Two near-miss incidents associated with clerical errors were experienced with group-specific blood transfusion. We conclude that urgent transfusion of uncrossmatched blood is safe, and that group O blood transfusion may be safer than group-specific blood transfusion in some emergency situations. Off-hour manuals based on the time required for blood preparation will avoid transfusion errors.

Adenoviral VEGF-A gene transfer induces angiogenesis and promotes bone formation in healing osseous tissues.

Vascular endothelial growth factor (VEGF) is a key regulator of angiogenesis. VEGF has been safely and efficiently applied to stimulate neovascularization in ischemic tissues in atherosclerotic patients. VEGF has an important role in bone repair by promoting angiogenesis and by stimulating major skeletal cell populations, chondrocytes, osteoblasts and osteoclasts. We studied the effect of VEGF-A on the recovery of bone drilling defects in rat femur delivered with first-generation adenoviral vector. The virus was injected into the muscle layer surrounding the bone defect made by drilling and the healing was followed for 1, 2, and 4 weeks. The VEGF effect was first demonstrated with an increased number of FVIII-related antigen-positive blood vessels in the defect area 1 week after the procedure. The proportional area of remaining reparative tissue was significantly reduced in the VEGF-treated animals 2 weeks after the injury suggesting favorable effect on bone healing. Increased periosteal cartilage was seen at the early phases of healing suggesting endochondral ossification. VEGF overexpression, however, completed the endochondral phase earlier compared with the control condition. Bone mineral content was enhanced in the VEGF-treated femurs measured with peripheral quantitative computed tomography at a 2-week time point. Our data confirm the important role of VEGF in bone healing. We show for the first time that adenoviral VEGF-A gene transfer may modify bone defect healing in a rodent model.

Mortality of hepatitis B surface antigen-positive blood donors in England and Wales.

Large population-based cohort studies in areas of high hepatitis B virus (HBV) prevalence have provided the evidence establishing hepatitis B surface antigen (HBsAg) carriage as a risk factor for hepatocellular carcinoma (HCC) and liver disease. Fewer studies have examined this in Western countries, where both HBV infection and carriage are less common and transmission patterns differ. This is the only prospective population-based study to examine this relationship in Europe. In all, 2681 male and 977 female blood donors in England and Wales, found to be HBsAg positive during routine blood-donation screening, were followed up from recruitment in 1970-1982 to December 1999 and their cause-specific mortality was analysed. This was compared with that of the general population of England and Wales. During a mean of 22 years of follow-up, 17.4% of the 420 deaths were due to HCC or liver disease. There were 20 deaths from HCC in male HBsAg carriers, representing a significantly high standardized mortality ratio (SMR) compared to the male population of England and Wales of 26 (SMR = 26.26; 95% CI: 16.04- 40.54). The HCC incidence rate in males was 33.5 per 100 000 person years and 4.4 per 100 000 person years in females. Men had 8.5 (SMR = 8.50; 95% CI: 6.25- 11.31) and women had 3.9 times the risk of death from liver disease (SMR = 3.89; 95% CI: 1.26-9.09). The risk of circulatory disease deaths was reduced in both males and females. There was a significant increased risk of non-Hodgkins lymphoma that was not apparent in the first decade of follow-up. The increased risk of HCC and liver disease in men fell with follow-up. Hepatitis B surface antigen carriage is a significant risk factor in England and Wales for both liver disease and HCC mortality. However, this risk has declined with duration of follow-up. This could be due to natural reversion to HBsAg negativity or as a result of treatment and avoidance of other risk factors. The increased risk of non-Hodgkins lymphoma seen in longer follow-up is likely to be related to HIV infection acquired subsequent to recruitment.

Strongly acidic electrolyzed water: valuable disinfectant of endoscopes

Background: Glutaraldehyde (GA) is currently considered to be the best disinfectant for endoscope disinfection. However, GA poses high risks for medical staff involved in the process and also to the environment. Strongly acidic electrolyzed water (SAEW) has been recently re‐evaluated for its potent bactericidal effect and environmental safety.Methods: Through the aspiration channel of the scopes, upper GI endoscopes and colonoscopes were experimentally contaminated with Pseudomonas aeruginosa, Mycobacterium avium and hepatitis B surface antigen positive blood. Four disinfection methods were tested: manual washing only, soaking in 3% GA for 5 and 10 min, and a 10‐s soak in SAEW with 50 or 100 mL of aspiration.Results: Direct plating culture was positive for Pseudomononas contamination after manual washing only (1/5) and after a 5‐min soak in 3% GA. Complete disinfection, confirmed by enrichment culture and polymerase chain reaction (PCR) of Pseudomonas and hepatitis B surface antigen positive blood on the contaminated upper GI endoscope was obtained after a 10‐min soak in GA and after using SAEW (0/5). Mycobacterum avium are rather resistant against SAEW as determined by broth culture and PCR (1/5).Conclusion: Strongly acidic electrolyzed water is a valuable disinfectant for endoscopes.

Inhibition of Hepatic Stellate Cell Proliferation and Activation by the Semisynthetic Analogue of Fumagillin TNP-470 in Rats

Proliferation and activation of hepatic stellate cells (HSCs) are critical steps for the development of postnecrotic fibrosis in the liver. The present study aimed to reveal the inhibitory effect of the semisynthetic analogue of fumagillin TNP-470 on these events for its possible use as an antifibrogenic agent. Rat models of carbon tetrachloride (CCl 4 )- and dimethylnitrosamine-induced hepatic fibrosis were used for an in vivo study. In both models, the fibrotic area was considerably decreased by concurrent repetitive subcutaneous injections of 30 mg/kg body weight of TNP-470. In CCl 4 -induced fibrosis, factor VIII-related antigen-positive blood vessels, desmin-, or α-smooth muscle actin (αSMA)-positive mesenchymal cells, bromodeoxyuridine (BrdU)-positive mesenchymal cells also decreased in number by treatment with TNP-470. In in vitro experiments, a supplement of 1,000 ng/mL TNP-470 suppressed BrdU incorporation and cyclins D1, D2, and E expression by cultured HSCs in the absence and/or presence of platelet-derived growth factor (PDGF). Expression of HSC activation markers, i.e., αSMA and PDGF receptor β, was also suppressed. The present results indicate that TNP-470 inhibits HSC proliferation by blocking the cell-cycle transition from G1 to S and HSC activation, and, as the consequence, prevents the progression of hepatic fibrosis, probably being coupled with its antiangiogenic effect. (Hepatology 2000;32:980-989.)

Detection of human cytomegalovirus-antigen in peripheral blood cells by using flow-cytometer

Prophlaxis of CMV-associated disease and/or early initiation of therapy is important in the management of patients with CMV infection. Recently, we developed the CMV antigenemia assay: a rapid and quantitative method based on the detection of CMC antigens in peripheral blood leukocytes by flow cytometry. Heparinized peripheral blood was obtained from healthy donors, bone marrow transplantation patients, patient with collagen disease and patients with adult T cell leukemia. To determine the phenotype of HCMV-infected mononuclear cells, the following phycoerythrin-conjugated mAb were used: CD8, CD15. These mAds were added to the whole blood and incubated. After the hemolysis, the cells were fixed with 4% paraformaldehyde and 0.3% NP-40. To determine the HCMV-infected cells, the following mAb were used in flow cytometry analysis: E13 (Chemicon Inc., Toyo) against HCMV IEA. As a secondary antibody, a FITC-conjugated goat anti-mouse IgG. In the bone marrow transplant patients, CMV-associated antigen was positive in their monocytes and polymorphocytes. In the patient with collagen disease, CMV-antigen was positive in their lymphocytes and monocytes. Our study demonstrates that the detection of CMV antigen-positive blood leukocytes by FACScan is a rapid and quantitative method and useful for the diagnosis and monitoring of CMV-associated CMV-associated disease. The CMV blood antigen assay by FACScan will facilitate the initiation of early treatment with ganciclovir of CMV-associated disease or the administration of prophylactic ganciclovir for preventing the disease.

The precore/core promoter mutant (T1762A1764) of hepatitis B virus: clinical significance and an easy method for detection.

Recently, a new hepatitis B virus (HBV) mutant with HBe antigen-negative phenotype has been characterized, in which one TATA box-like motif of the precore/core promoter had degenerated: most frequently by both A-->T and G-->A mutations at positions 1762 and 1764, respectively. The clinical significance of this mutant is as yet unknown. In our present study, the T1762 A1764 mutant was sought in sera from HBV-infected blood donors and chronic liver disease patients by directly sequencing a PCR-amplified region of HBVDNA. Also, because the A1764 mutation generates a Sau3AI cleavage site (GGTC-->GATC), we digested the PCR products with Sau3AI to see if cleavage would occur at this specific site. Our results mostly corroborated the earlier report but we found a higher-than-predicted frequency of HBe antigen-positive blood donors positive for the mutant (22%). The titres of HBe antigen in these mutant-positive sera were slightly decreased compared to the titres in wild-type HBV infection. In addition, these blood donors had relatively high (though within the normal range) serum alanine aminotransferase (ALT) levels, suggesting that the T1762 A1764 mutation could be used as a sensitive laboratory marker for insidious hepatitis in these otherwise 'asymptomatic' carriers. The Sau3AI assay, which is much more convenient than sequencing, was shown to be useful for the detection of the T1762 A1764 mutant in an extensive number of clinical samples.

Cytomegalovirus (CMV) antigenaemia for rapid diagnosis and monitoring of CMV‐associated disease after bone marrow transplantation

A technique for the rapid detection of cytomegalovirus (CMV) antigen-positive blood leucocytes (CMV antigenaemia) was evaluated in 15 marrow transplant patients as a means of diagnosis and for monitoring CMV-associated disease. CMV antigenaemia was determined by direct immunoperoxidase staining of leucocytes with a peroxidase-labelled monoclonal antibody, HRP-C7, which binds an immediate-early antigen of human CMV. CMV antigenaemia occurred in 7/15 marrow transplant patients (47%) and was initially detected between 4 and 6 weeks after transplantation. CMV-associated diseases developed in 3/15 patients (20%). All patients with CMV-associated disease had a relatively large number of CMV antigen-positive leucocytes, exceeding 10 per 50,000 white blood cells (WBCs). In the remaining 12 patients, CMV antigen-positive leucocytes were less than 10 per 50,000 WBCs or were undetectable. CMV-associated disease did not develop in these patients during the period of monitoring. CMV antigen-positive leucocytes were detected more frequently in patients who developed acute graft-versus-host disease (GVHD) or haemorrhagic cystitis than in those without such complications. CMV antigens were detectable from 1 to 4 weeks before the onset of CMV-associated disease which allowed initiation of ganciclovir treatment at an early stage. The degree of CMV antigenaemia paralleled the clinical symptoms and signs, higher degrees of antigenaemia being associated with more significant disease. Thus, the detection of CMV antigen-positive blood leucocytes is useful for the diagnosis and monitoring of CMV-associated disease following bone marrow transplantation.

Lack of clinical significance of "enzyme-only" red cell alloantibodies.

In a retrospective study on samples from 10,000 recently transfused patients, 35 samples were found to contain an antibody that reacted with ficin-treated red cells but was not demonstrable by low-ionic-strength saline solution and indirect antiglobulin test (LISS-IAT). In those 35 patients, the specificity of the antibody was such that each patient would have been transfused with antigen-negative blood had the antibody reacted in LISS-IAT. Tests on red cells from the units already transfused showed that 19 patients had among them received, by chance, 32 antigen-positive and 74 antigen-negative units. The remaining 16 patients had among them received 57 units that were, again by chance, all antigen negative. One patient given antigen-positive blood suffered a delayed transfusion reaction; in two others the antibodies became LISS-IAT active after transfusion. However, similar changes to the LISS-IAT-active state were seen with two antibodies of patients given only antigen-negative blood. Also found in the 10,000 patients were 28 clinically insignificant antibodies, 77 sera in which the antibody was too weak to identify, and 216 autoantibodies that reacted only with ficin-treated red cells. These data support a belief, generally held in the United States but not necessarily elsewhere, that the use of protease-treated red cells for routine pretransfusion tests creates far more work than the accrued benefits justify.