Antiglobulin Test Research Articles

Cord Direct Antiglobulin Test for Predicting the Need for Phototherapy in Neonates with ABO Incompatibility: A Prospective Cohort Study

Introduction: The Direct Antiglobulin Test (DAT) is a screening technique used to detect Immunoglobulin G (IgG) antierythrocyte antibodies on foetal Red Blood Cells (RBCs). Universal bilirubin screening decreases the incidence of severe hyperbilirubinemia. The diagnostic efficacy of DAT in cases of ABO incompatibility is not well established, in contrast to Rh incompatibility. Aim: To evaluate the diagnostic ability of routine cord blood DAT in neonates ≥35 weeks of gestation with ABO incompatibility in predicting the requirement for phototherapy. Additionally, to compare the diagnostic ability between two methods of DAT estimation, namely the semiagglutination gel method and the conventional tube method. Materials and Methods: This prospective cohort study was conducted from September 2022 to March 2024 at SRM Medical College Hospital and Research Centre, Chennai, Tamil Nadu, India. Following the acquisition of informed consent, cord blood samples were collected from 934 neonates delivered after 35 weeks of completed gestation to mothers with O blood type. In cases of ABO incompatibility (523 neonates), DAT was performed using both the manual tube method and the automated semiagglutination gel method in the blood bank. Serum bilirubin levels, complete blood counts with peripheral smear analysis and reticulocyte counts were performed on day 3 or earlier, as clinically indicated. The neonates were treated in accordance with the American Academy of Paediatrics (AAP) 2004 recommendations on phototherapy. Babies were monitored for jaundice until 14 days of age. Results: Out of 3,092 eligible deliveries, 523 babies with ABO incompatibility were enrolled. The baseline characteristics were comparable. Bilirubin levels in DAT-positive and DAT-negative neonates were 14.60±3.58 mg% and 13.18±3.16 mg% (p-value <0.001), respectively. The adjusted odds ratio of positive DAT for both methods in predicting the requirement for phototherapy was 3.21 (95% CI 2.1-5.8). The sensitivity, specificity, positive likelihood ratio and negative likelihood ratio of the tube method were 37.5%, 85.16%, 2.53, and 0.73, respectively, while for the gel method they were 37.08%, 86.57%, 2.76, and 0.73, respectively. Conclusion: In cases of ABO incompatibility, neonates with positive cord DAT had higher odds of requiring phototherapy; however, a high negative likelihood ratio indicates that additional factors may be involved in hyperbilirubinemia that necessitate phototherapy.

Pregnancy Favism and Severe Hemolytic Anemia in a Patient with Glucose-6-Phosphate Dehydrogenase (G6PD) Deficiency: A Case Report

Introduction: Favism is a genetic disease-causing hemolytic anemia in Mexico, primarily due to glucose-6-phosphate dehydrogenase (G6PD) deficiency. Factors triggering favism include infections, pregnancy, certain drugs, and eating beans. The disease is inherited recessively and is more common in men. Diagnosing and managing favism during pregnancy is crucial, as it can lead to severe hemolytic anemia and neonatal jaundice. Case presentation: A female patient, aged 18, presented with severe anemia and thrombocytopenia during her 20-week pregnancy. During pregnancy, the patient experienced jaundice and anemia at 21.5 weeks of gestation. Despite not presenting evidence of bleeding, transfusions were performed due to critical hemoglobin levels. G6PD deficiency was confirmed at 9 days of hospital admission, indicating severe hemolytic anemia. Clinical discussion: The hemolytic crisis in a pregnant woman with favism was triggered by pregnancy and repeated infections of candidiasis. Clinical symptoms of hemolysis were confirmed through G6PD and Coombs tests, and blood transfusion was recommended to prevent complications. The patient's hemoglobin was monitored for signs of active intravascular hemolysis, and a folic acid tablet was recommended during the crisis. Post-crisis management involved periodic controls of hemoglobin, hematocrit, lactate dehydrogenase, and bilirubin. Conclusion: Favism, a prevalent condition in Mexico, should be considered in patients with a family history of hemolytic anemia or compatible symptoms. Early diagnosis and proper management are crucial to prevent complications in the mother and fetus. A multidisciplinary approach, genetic counseling, and medical education are essential for effective management.

Direct antiglobulin test for the prediction of neonatal hyperbilirubinemia needing an intervention: a systematic review and diagnostic test accuracy meta-analysis

ImportanceThe direct antiglobulin test (DAT) is commonly used as a screening test for predicting significant neonatal hyperbilirubinemia requiring intervention. However, evidence for this approach is limited.ObjectiveThe aim of this study was to evaluate the diagnostic utility of DAT in predicting the need for phototherapy and double volume exchange transfusion (DVET) in neonates with ABO and Rhesus (Rh) incompatibility conditions.MethodsMEDLINE, Embase, CENTRAL, CINAHL, and Web of Science were searched from inception until 1 February 2024. Randomized controlled trials (RCTs) and non-RCTs were eligible for inclusion. Two reviewers screened the titles and abstracts blinded to each other. A Bayesian bivariate random-effects model was employed for the diagnostic test accuracy meta-analyses. Risk of bias was assessed using Quality Assessment for Studies of Diagnostic Accuracy 2 and certainty of evidence (CoE) was adjudged according to the Grading of Recommendations, Assessment, Development, and Evaluations (GRADE) guidelines.ResultsIn total, 53 studies were included in the systematic review and 28 were synthesized in the meta-analysis. For the need for phototherapy outcome, the pooled sensitivity [95% credible interval (CrI)] and specificity (95% CrI) of DAT in ABO incompatibility (18 studies, n = 10,110) were 56.1% (44.5%–67.8%) and 83.6% (71.6%–90.8%). For Rh incompatibility (three studies, n = 491), the sensitivity and specificity were 40.4% (12.2%–81.7%) and 89.9% (72.7%–94.6%). The CoE was predominantly low. For the need for DVET outcome, the pooled sensitivity and specificity of DAT in ABO incompatibility (three studies, n = 2,652) were 83.6% (35.8%–99.6%) and 74.5% (40.3%–92.7%). For Rh incompatibility (two studies, n = 240), the sensitivity and specificity were 80.3% (34.2%–97.3%) and 68.0% (25.3%–92.1%). The CoE was predominantly very low.ConclusionIn ABO and Rh incompatibility, DAT probably has moderate specificity and low sensitivity for predicting the need for phototherapy. For DVET, though DAT is possibly a better predictor due to its acceptable sensitivity, the predictive interval was wide. Thus, we do not suggest the routine use of DAT screening to predict the need for phototherapy and DVET. However, it may be used as a second-tier investigation for risk stratification of high-risk neonates.Systematic Review Registrationhttps://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42022297785, PROSPERO (CRD42022297785).

A whistleblower direct antiglobulin test in chronic kidney disease: An interesting case of occult autoimmune hemolytic anemia in an undiagnosed lymphoma with pure red cell aplasia

Abstract A 67-year-old male, presented with urosepsis, severe anemia, advancing azotemia requiring blood transfusion. Patient grouped as O RhD positive. Antibody screening was pan-positive (4+). Similar strong pan-positivity seen on antibody identification with extended cell panel. DAT was 4+ positive for IgG and negative for complement C3. Repeated identification with eluate also showed pan-positive (4+) reaction. Serum haptoglobin markedly reduced. Bone marrow aspirate suggested pure red cell aplasia with suspicion of parvovirus-related changes. Bone marrow biopsy suggested T-cell lymphoproliferative disorder. Immunophenotyping elucidated a suspicious clone of CD7-negative T-cells with CD4 restriction. Histopathological examination revealed angioimmunoblastic T-cell lymphoma (AITL) with bystander Epstein–Barr virus-driven large B-cells. Owing to rarity and nonspecific symptoms, diagnosis of AITL is often challenging. Antibody screen acted as a whistleblower for an undiagnosed aggressive lymphoma. Thus, it will be prudent to look beyond the obvious in complex cases, and a detailed workup will help put together all parts of the jigsaw puzzle.

The Prevalence of Peripheral Erythrophagocytosis in Pediatric Immune-Mediated Hemolytic Anemia.

Peripheral erythrophagocytosis appears to be a unique sign of acquired immune-mediated hemolytic anemia. It is said to be rare but its prevalence among patients with autoimmune hemolytic anemia has not been studied. In this retrospective study from July 2014 to June 2024, the clinical and laboratory features, treatment and outcomes of children diagnosed with autoimmune hemolytic anemia were described. The prevalence of peripheral erythrophagocytosis was compared to a group of children with hereditary spherocytosis at the time of first diagnosis seen in the same period. Twelve consecutive children with autoimmune hemolytic anemia were included. There were four female patients. The mean age was 6.7 (range 0.8 to 16.6) years. The mean hemoglobin was 6.0 (range 2.5 to 8.1) g/dL. Seven patients were positive by a direct antiglobulin test, three were positive with cold agglutinins and two were positive on both tests. In seven cases, an acute infection appeared to be the precipitating factor. Mycoplasma pneumoniae infection was documented in three and suspected in another two cases. Peripheral erythrophagocytosis was present in five cases (42%) but was not found at diagnosis in any of the 16 cases of hereditary spherocytosis (p = 0.0081). Six children had pre-existing diseases, including two with hereditary hemolytic anemia. Peripheral erythrophagocytosis is a relatively common and characteristic finding in pediatric autoimmune hemolytic anemia and should be actively looked for in the evaluation of acute hemolysis, including in children with pre-existing hereditary hemolytic disorders.

The Discovery of New Antibody in Autoimmune Disease Using a Novel Approach of Coombs Test Based on Flow Cytometry Method.

The objective of this study was to investigate the antibody to RBC in autoimmune disease patients with ANA using sensitive Coombs test based on flow cytometry method. Antinuclear antibodies (ANA) of autoimmune disease patients were added to red blood cells (RBCs) of blood group O. At the same time, healthy individuals' serums were also checked. The sample tubes were incubated for 30 min at 37°C. After incubation, each sample was analyzed on flow cytometry. The agglutination of antinuclear antibodies in autoimmune patients was observed, while in healthy people, there was no agglutination between RBCs and serum. A significant difference was found between the disease group and healthy group (p < 0.0001) showing that a statistical analysis was conducted to compare the presence of ANA agglutination between the two groups. The reported p-value of less than 0.0001 shows that the observed difference is highly significant. The serum stability test conducted over ten consecutive days demonstrated a CV of 6.90% in the test results, indicating favorable stability. In conclusion, this study emphasizes the effectiveness of flow cytometry as a valuable tool for detecting RBC-bound antibodies and new antibodies in autoimmune disease patients. Its high sensitivity and accuracy have the potential to greatly improve diagnostic capabilities in clinical laboratories.

Sovleplenib in patients with primary or secondary warm autoimmune haemolytic anaemia: results from phase 2 of a randomised, double-blind, placebo-controlled, phase 2/3 study.

Spleen tyrosine kinase inhibitors are potential treatment options for warm autoimmune haemolytic anaemia. This study aimed to assess the preliminary efficacy and safety of sovleplenib-an oral spleen tyrosine kinase inhibitor-in patients with warm autoimmune haemolytic anaemia in China. Here we report on the phase 2 results. This randomised, double-blind, placebo-controlled, phase 2 part from the phase 2/3 study was conducted at 13 centres in China. Eligible patients, aged 18-75 years, with an Eastern Cooperative Oncology Group (ECOG) performance status of no more than 2, had primary or secondary warm autoimmune haemolytic anaemia (stable underlying disease not requiring drug intervention) with no response to previous glucocorticoid treatment, haemoglobin of less than 100 g/L with active haemolysis, and a positive direct antiglobulin test. The study comprised two periods; patients were randomly assigned (3:1) to receive sovleplenib or placebo at 300 mg orally once a day in the 8-week double-blind period. Upon completion, all patients entered an open-label treatment period for at least 16 weeks and received sovleplenib 300 mg once a day until 24 weeks after the last patient was randomly assigned. The primary endpoint for phase 2 of the trial was overall haemoglobin response rate (haemoglobin ≥100 g/L with an increase of ≥20 g/L from baseline at least once, and haemoglobin not affected by rescue therapy, such as red blood cell transfusions, intravenous immunoglobulin, and glucocorticoids) by week 24. Efficacy analyses in the 0-8 week double-blind period included all patients who were randomly assigned, analysed by intention-to-treat. Safety analysis in the double-blind period included patients in the intention-to-treat population who received at least one dose of the study medication. This phase 2/3 study is registered with ClinicalTrials.gov, NCT05535933, and the phase 3 part is ongoing. Between Sept 26, 2022, and May 9, 2023, 34 patients were screened and 21 patients (four [19%] male and 17 [81%] female) were enrolled in the study and randomly assigned to receive either sovleplenib (n=16) or placebo (n=5). All 21 patients completed the 0-8-week double-blind treatment and entered the open-label treatment period. The overall haemoglobin response rate was 67% (14 of 21 patients) by week 24, and durable haemoglobin response rate was 48% (ten of 21 patients) by week 24. During the 0-8-week double-blind treatment, 13 (81%) of 16 patients in the sovleplenib group versus five (100%) of five patients taking placebo reported treatment-emergent adverse events (TEAEs), and four (25%) of 16 patients versus four (80%) of five patients reported grade 3 adverse events. Although all 21 patients had a TEAE during the 24-week treatment with sovleplenib, only seven (33%) patients had grade 3 events. The most common grade 3 TEAE was anaemia (four [19%] patients), which was not related to treatment. There were no grade 4 or 5 TEAEs. Sovleplenib treatment achieved an encouraging overall haemoglobin response in Chinese patients with warm autoimmune haemolytic anaemia and was well tolerated. The phase 3 part of the study (ESLIM-02) is currently ongoing to further substantiate the efficacy and safety of sovleplenib in this setting. HUTCHMED.

Systemic lupus erythematosus in an adolescent male: A rare case presentation

Abstract Pediatric systemic lupus erythematosus (pSLE) is a rare but severe autoimmune condition with multisystem involvement, disproportionately affecting females. Here, we report the case of a 15-year-old male presenting with polyarthralgia, fatigue, and intermittent fever, accompanied by photosensitive skin rash, oral ulcers, and nonproductive cough. Examination revealed a malar rash sparing the nasolabial fold, hard palate ulcers, and a palpable purpuric rash. Initial investigations demonstrated bicytopenia, elevated inflammatory markers, and liver enzyme derangements. Autoimmune workup confirmed pSLE per Systemic Lupus International Collaborating Clinics criteria, with positive anti-double-stranded DNA, anti-Smith antibodies, low complement levels, and direct Coombs test positivity. Radiological findings included minimal pleural effusion and mild ileitis. The patient was managed with pulse methylprednisolone, oral prednisolone, hydroxychloroquine, and cyclophosphamide, resulting in clinical improvement. This case underscores the diagnostic challenges of pSLE in male adolescents and highlights atypical manifestations such as scalp lesions mimicking tinea capitis. Early recognition and timely intervention are crucial for the favorable outcomes.

Risk prediction of new-onset thrombocytopenia in patients with systemic lupus erythematosus: a multicenter prospective cohort study based on Chinese SLE treatment and research group (CSTAR) registry

BackgroundThrombocytopenia (TP) is a hematological manifestation of systemic lupus erythematosus (SLE) and is associated with unfavorable prognostic outcomes. This study aimed to develop a risk prediction model for new-onset TP in SLE patients.MethodsBased on the multicenter prospective Chinese SLE Treatment and Research Group (CSTAR) registry, newly diagnosed SLE patients without TP at registration were enrolled. The least absolute shrinkage and selection operator (LASSO) method was used for variable selection. The final model was developed using multivariate Cox regression and displayed as a nomogram. Internal validation was achieved using enhanced Bootstrap resampling.ResultsDuring follow-up, thrombocytopenia developed in 80 (3.52%) of 2270 lupus patients. The final risk prediction model incorporated six predictors: baseline SDI score ≥ 1 (HR 2.207, 95% CI 1.350–3.609, p = 0.002), hemolytic anemia (HR 1.953, 95% CI 1.017–3.750, p = 0.044), low complement level (HR 2.351, 95% CI 1.004–5.505, p = 0.049), positive anti-β2GPI antibody (HR 1.805, 95% CI 1.084–3.004, p = 0.024), positive Coombs test (HR 1.878, 95% CI 1.123–3.141, p = 0.017), and positive anti-histone antibody (HR 1.595, 95% CI 1.017–2.587, p = 0.059). The model’s performance was indicated by C-index values for risk prediction at one, two, and three years, which were 0.741 (0.660–0.823), 0.730 (0.655–0.805), and 0.710 (0.643–0.777), respectively; and Brier scores of 0.018 (0.012–0.024), 0.025 (0.017–0.032), and 0.037 (0.027–0.046), respectively. Calibration curves were drawn and situated near the diagonal line.ConclusionsThis study developed the first risk prediction model for TP onset in lupus patients. Patients with baseline organ damage, hemolytic anemia, low complement, positive anti-histone antibody, positive anti-β2GPI antibody, or positive Coombs test were identified as being at high risk for thrombocytopenia and require further clinical attention.

Alloantibody Identification: The Importance of Temperature, Strength Reaction and Enzymes-A Practical Approach.

Red blood cell (RBC) alloimmunization and antibodies formation against non-self antigens on red cells may occur after blood transfusion, pregnancies or other exposures. The RBC alloimmunization rate varies from 2% to 6% according to recent studies. The antibody screen is performed to identify or confirm the presence of antibodies in patient's serum or plasma, as a preoperative or pretransfusion test. The antibody identification process and major crossmatch are critical steps of risk management in transfusion medicine. The aim of this article is to describe a flow chart of the antibody identification. I report three educational examples of case studies associated with the negative direct antiglobulin test and clinically significant single and multiple alloantibodies using the gel method, Anti-M, Anti-c and Anti-E, Anti-Jka and Anti-s. Furthermore, I provide a critical analysis of the current literature on the topic. The flow chart of the antibody identification may simplify the process and possibly reduce errors in routine workflow.

RhD-negative red blood cells can be saved during liver transplantation in RhD-negative patients due to low risk of alloimmunization against RhD.

Transfusion demand is high in liver transplantation (LT), and thus RhD-positive (RhD+) red blood cell concentrates (RBCs) are sometimes given to RhD-negative (RhD-) patients. Due to immunosuppression, these patients rarely produce anti-D. We investigated the rate of anti-D formation in RhD- patients undergoing LT who were transfused with RhD+ RBCs as well as the number of transfused RhD- and RhD+ RBCs. RhD-type and transfusion history of all patients undergoing LT between 2010 and 2023 were reviewed retrospectively. In RhD- patients, who received RhD+ RBCs, the results of antibody screening test (indirect antiglobulin test and with papain-treated test cells) and direct antiglobulin test were evaluated. Five hundred and twenty-seven patients underwent 576 LT. Eighty-seven patients were RhD-, of whom 42 were transfused with RhD+ RBCs. In 34 of them, an antibody screening test result was available more than two weeks after the last RhD+ RBCs transfusion. In two of them, a transient, weak anti-D antibody was detectable, which disappeared in the further course. Overall, 1352 RBCs were transfused to the 87 RhD- patients, 543 of those were RhD+. Most RhD+ RBCs were provided to men and elder women. Transient weak anti-D occurred in two RhD- male patients during LT after transfusion of RhD+ RBCs without evidence for a hemolytic transfusion reaction. To save stocks of RhD- RBCs, early transfusion of RhD+ RBCs to RhD- men and women beyond the childbearing age should be considered during LT.

Blood Cross Matching Without Anti-Human Globulin (AHG) and Bovine Serum: A New Interest for an Old Idea

Introduction Transfusion medicine promotes the safety of blood transfusions by rigorously testing to eliminate risks of infection and hemolytic. The efficacy (to correct and identify antibodies) of a modified cross-matching method that excludes the use of anti-human globulin (AHG) bovine serum with that of the immediate spin crossmatch was assessed. Material and Methods This multi-center study was performed at two medical centers in Iran and Iraq. Over seven years, consecutive participants received two different blood cross-matching methods: one with AHG and bovine serum and another without it. Results The study included 31240 participants. About 18526 (59.3%) were males and 12714 (40.7%) were females. The ages of participants ranged from 23 to 57 years, with the average age of 39 years. Only 45 (0.14%) participants blood cross-match incompatible in both spin cross-match and cross-match without AHG, bovine serum the same result. The study found that all detected antibodies correlated with potential blood incompatibility, and there remained 100% safety between the two testing methods. Conclusion Omitting AHG and bovine serum in cross-matching might be safe. At least it can be used in emergency situations and in resource-limited settings.

Clinical Characteristics and Prognosis of Hemolytic Disease of the Newborn Caused by Irregular Antibodies: A 13-Year Retrospective Analysis.

The clinical characteristics and outcomes of hemolytic disease of the newborn (HDN) caused by irregular antibodies remain unclear. Herein, we analyzed the clinical features and prognosis of HDN. Children admitted to our institution between June 2009 and December 2022 with a definite diagnosis of HDN were evaluated. Patients with irregular antibodies were matched in a 1:3 ratio to those with ABO incompatibility. Children with confirmed Rh-incompatibility hemolytic disease were divided into the RhD subgroup (hemolysis induced by Rh anti-D) and the non-RhD group (hemolysis induced by other Rh antibodies). The irregular antibody and ABO incompatibility group included 32 and 96 patients, respectively. Compared to the ABO incompatibility group, the irregular antibody group showed earlier jaundice; higher incidence of liver and spleen enlargement and anemia; higher direct antiglobulin test (DAT) positivity; earlier and more severe anemia; higher rates of enhanced phototherapy, blood transfusion, and blood exchange; and longer hospital stay (all p < 0.05). Compared to the non-RhD group, the RhD subgroup showed an earlier occurrence of jaundice and a higher incidence of liver and spleen enlargement (both p < 0.05). The multiple irregular antibody subgroup further showed earlier occurrence of jaundice and a higher rate of enhanced phototherapy, blood transfusion, and blood exchange than the single-antibody group (both p < 0.05). HDN caused by irregular red blood cell antibodies is rare, but clinical manifestations are serious. It is important to pay attention to the screening of irregular antibodies during pregnancy, to strengthen monitoring, and to provide intrauterine treatment and early intervention as necessary.

A Rare Case of Warm Autoimmune Hemolytic Anemia and Massive Splenomegaly in Uncontrolled Grave's Disease

Background: Autoimmune hemolytic anemia (AIHA) occurs when a person's immune system produces autoantibodies that attack their red blood cells, leading to their destruction. Warm AIHA, the most common form of AIHA, is caused by antibodies such as IgG that are active at body temperature. Warm AIHA can be primary or associated with underlying conditions like autoimmune disorders, lymphoproliferative disorders, immunodeficiencies, and infections, due to some form of immune system disruption. Clinical signs include anemia, jaundice, and mild to moderate splenomegaly. Case Presentation: A 36-year-old man with a history of Graves' disease (GD) (on methimazole) presented with jaundice. Despite being on methimazole, his thyroid levels were uncontrolled due to medication non-adherence, so his outpatient methimazole dose had been increased to 40 mg. Initial labs were notable for the following: a white blood cell count of 1.5 k/mm3, hemoglobin (Hgb) of 5.0 g/dL, platelet count of 78 k/mm3, total bilirubin of 13.1 mg/dL, reticulocyte percentage of 13.1%, lactate dehydrogenase (LDH) of 304 U/L, undetectable haptoglobin, absolute neutrophil count (ANC) of 0.95 k/mm3, thyroid stimulating hormone &amp;lt;0.01 mIU/L and a free T4 of 3.77 ng/dL. CT scan of the abdomen pelvis with contrast showed splenomegaly measuring 24.4 cm. Some reticulocytes were noted on the peripheral smear. The patient was diagnosed with AIHA, with warm antibodies noted on the Coombs test. In terms of workup for the etiology of his AIHA, the following infectious workup was negative: blood parasite smear, HIV, and Lyme enzyme immunoassay. Bone marrow (BM) biopsy flow cytometry showed no immunophenotypic evidence of acute leukemia, non-Hodgkin lymphoma, or high-grade myelodysplasia. He was aggressively transfused with 5 units of PRBCs and started on 1 mg/kg of prednisone daily. He was started on filgrastim 300 mcg 2 times a week for his leukopenia. Post these treatments his hemolysis labs improved to Hgb of 10.4 g/dL, reticulocyte percentage of 4.02%, total bilirubin of 3.1mg/dL, LDH of 141 U/L and haptoglobin of 78 mg/dL. His methimazole was held inpatient due to low ANC count. A trial of lithium was started for GD which led to limited improvement. The patient was discharged on daily prednisone and planned weekly rituximab infusions for AIHA. The patient was discharged without an anti-thyroid agent, with plans for close follow-up outpatient for thyroidectomy vs radioiodine ablation. Conclusion: The patient's severe Warm AIHA appeared acute, however the massive splenomegaly indicated a chronic low-level hemolytic process. Because the patient did not have any diagnostic findings consistent with hematologic malignancy or infection, and given the context of his recent history, it is reasonable to suspect that this presentation can be attributed to uncontrolled GD. Although the exact pathophysiology of the association is still unclear, the autoimmune nature of GD and hyperthyroidism are thought to contribute to the hemolysis. Warm AIHA is the rarest hematologic manifestation of GD, and literature on the topic has been limited to single case reports. Our case is also unique in that the Warm AIHA was seen in a patient with established GD, and not as an initial presentation of the disorder. To our knowledge, this is also the first case of Warm AIHA with this severe degree of splenomegaly, which can only be explained by the chronic nature of the patient's GD. The response to treatment for AIHA suggests an autoimmune nature of the condition, but the underlying GD did not respond to medical therapy. Overall, this case adds to the literature, as a unique hematologic presentation of an uncontrolled autoimmune disorder.

Using Soluble CD38 to Overcome Daratumumab Interference in Pretransfusion Compatibility Testing

Background: CD38 is a protein highly expressed on myeloma (MM) cells that has been shown to be an effective target antigen for monoclonal antibody therapies, so treatment with anti-CD38 monoclonal antibodies is a first line therapy for patients with MM. Although CD38 is weaker expressed on erythocytes anti-CD38 binds to CD38 on reagent RBCs and cause panreactivity in vitro and subsequent false positive reactions in indirect antiglobulin tests (IAT), antibody detection (screening) tests, antibody identification panels, and anti-human globulin (AHG) crossmatches. These findings suggest that the soluble CD38 method could improve transfusion safety for patients receiving anti-CD38 therapy, particularly in urgent clinical settings This study aims to evaluate the effectiveness of a new soluble CD38-based method in mitigating Daratumumab interference during pretransfusion compatibility testing." Methods: We evaluated the Grifols sCD38 method in 20 patients and compared it with the DTT method Results: In our experience the sCD38 method reduced testing time from 150 to 50 minutes and demonstrated 100% efficacy, compared to 90% with DTT. The sCD38 effectively mitigated the interference caused by anti-CD38 antibodies in 10 (100% efficacy) of patient samples tested while DTT was successful in only 9/10 (90% efficacy); no interference was observed in patients presenting anti erythrocyte antibodies. Moreover, there was no negative influence on DTT sensitive blood group systems such as KEL upon sCD38 treatment. Conclusions: In our Laboratory, in the year 2023, we processed 129 patients treated with anti-CD38. Therefore the reduction from 150 to 50 minutes in the time needed to perform tests for mitigation of anti-CD38 interference appears to be relevant with a recovery of approximately 210 technical-hours per year. Another highly appreciated operational aspect was the possibility of treating the patient's plasma and perform tests using automatic instruments (Erytra Grifols) available in our laboratory. In the evaluation of this new method, we did not observe failures in the mitigation of anti-CD38 interference. Furthermore, the results obtained in the samples that presented allo or auto antibodies were not affected by the treatment with sCD38. In our experience Grifols sCD38 assay is straightforward and quick to perform ant it is superior to DTT treatment in the mitigation of anti-CD38 antibody interference in MM patients treated with Daratumumab.

Autoimmune haemolytic anaemias.

Adult autoimmune haemolytic anaemias (AIHAs) include different subtypes of a rare autoimmune disease in which autoantibodies targeting autoantigens expressed on the membrane of autologous red blood cells (RBCs) are produced, leading to their accelerated destruction. In the presence of haemolytic anaemia, the direct antiglobulin test is the cornerstone of AIHA diagnosis. AIHAs are classified according to the isotype and the thermal optimum of the autoantibody into warm (wAIHAs), cold and mixed AIHAs. wAIHAs, the most frequent type of AIHAs, are associated with underlying conditions in ~50% of cases. In wAIHA, IgG autoantibody reacts with autologous RBCs at 37 °C, leading to antibody-dependent cell-mediated cytotoxicity and increased phagocytosis of RBCs in the spleen. Cold AIHAs include cold agglutinin disease (CAD) and cold agglutinin syndrome (CAS) when there is an underlying condition. CAD and cold agglutinin syndrome are IgM cold antibody-driven AIHAs characterized by classical complement pathway-mediated haemolysis. The management of wAIHAs has long been based around corticosteroids and splenectomy and on symptomatic measures and non-specific cytotoxic agents for CAD. Rituximab and the development of complement inhibitors, such as the anti-C1s antibody sutimlimab, have changed the therapeutic landscape of AIHAs, and new promising targeted therapies are under investigation.

Protocolo diagnóstico de las anemias hemolíticas

Hemolytic anemias are defined by the premature destruction of circulating red blood cells. They can be classified as congenital or acquired and most are of immune origin. Diagnosis requires a comprehensive medical history, including a personal and family history; use of drugs or toxins; and a series of laboratory tests. Test findings predominantly show a marked decrease in hemoglobin; an increase in reticulocytes, bilirubin (indirect fraction), and lactate dehydrogenase; and a decrease in haptoglobin. Finally, a direct antiglobulin test (direct Coombs test) and a peripheral blood smear can be very useful in indicating the possible cause of hemolytic anemia.

Oxidised low-density lipoprotein and antibodies against oxidised low-density lipoprotein in patients with antiphospholipid syndrome.

Recurrent thrombosis is one of the main clinical features of antiphospholipid syndrome (APS), and recent studies revealed that APS shares similar pathophysiological mechanisms with atherosclerosis. Oxidised low-density lipoprotein (OxLDL) and antibodies against OxLDL (anti-OxLDL) are involved in the development of atherosclerosis. This study aims to investigate the clinical significance of OxLDL and anti-OxLDL in APS patients. One hundred and seventy APS patients and 39 healthy controls (HC) were enrolled. Clinical and laboratory data were collected from Clinical Data Center of Peking University People's Hospital. OxLDL and anti-OxLDL were detected using enzyme-linked immunosorbent assay. Among the 170 APS patients, 106 had isolated thrombotic APS. Compared with HC, APS patients exhibited higher titres of OxLDL [413.86 (220.11-853.67) ng/mL vs. 45.54 (0-105.98) ng/mL, p<0.001] and anti-OxLDL [107.62 (75.68-174.18) U/L vs. 44.13 (18.44-79.76) U/L, p<0.001]. Also, APS patients exhibited a higher positivity rate for OxLDL (88.2% vs. 5.1%, p<0.001) and anti-OxLDL (84.1% vs. 36.5%, p<0.001) compared to HC. APS patients with elevated levels of OxLDL had a higher rate of LAC positivity (68.0% vs. 45.0%, p=0.042). Furthermore, APS patients with positive anti-OxLDL demonstrated a higher occurrence of venous thrombosis (46.2% vs. 18.5%, p=0.008) and a lower rate of Coomb's test positivity (52.6% vs. 76.2%, p=0.049). Multivariate logistic regression revealed that anti-OxLDL positivity (OR 12.424, 95%CI 1.108-139.330, p=0.041) were risk factors for venous thrombotic APS. This study indicates that the presence of anti-OxLDL may serve as potential markers for venous thrombosis in APS patients. OxLDL and anti-OxLDL may function as valuable biomarkers for monitoring APS.

P39 Primary antiphospholipid syndrome before development of severe systemic lupus erythematosus – a case report

Abstract Introduction Antiphospholipid syndrome (APS) has long been associated with systemic lupus erythematosus (SLE). Graham Hughes first reported APS in 1983 in cohorts of SLE patients with recurrent thrombosis. We present a case of a young man who developed severe manifestations of SLE 7 years after the diagnosis of primary APS and we look at the literature for predictors in primary APS to develop into SLE. Case description A 28 year old Caucasian male with Turkish ancestry was diagnosed with triple positive primary APS at the age of 21 when he presented with bilateral pulmonary embolisms. He was stable on warfarin with target INR between 3-4. He had been having occasional mouth ulcers, lethargy, mild paraesthesia of his feet and headaches for 2 months. He then presented in February 2024 with short history of pleuritic chest pain and marked breathlessness. Initial cross sectioning imaging did not reveal pulmonary embolism. A week later, echocardiogram demonstrated large global pericardial effusion and small pleural effusions. Ejection fraction was preserved. Interval MRI scan showed pericarditis without evidence of myocarditis. Almost 600 mls of heavily blood stained pericardial fluid was drained. Analysis of the pericardial fluid showed inflammatory processes with macrophages, neutrophil and lymphocytes. Culture was negative including for tuberculosis and cytology negative for malignancy. Immunology revealed ANA 1:160, ENA normal, dsDNA was initially negative but upon interval testing was high at 27.5 IU/mL, IgG 13.1 g/L, C3 1.2 g/L, C4 &amp;lt;0.1 g/L, CRP 143, ESR 30. He had anaemia, lymphopenia and positive direct antiglobulin test. ANCA and rheumatoid factor were unremarkable. Renal and joints were unaffected. Due to his Turkish ancestry, he was extensively investigated for the possibility of Behcet’s with normal femoral vein ultrasound and negative HLA-B51 genotype. His manifestations fulfilled the SLICC criteria to be diagnosed with SLE. His response to intravenous methylprednisolone infusions and subsequent oral steroid was marked. He was initiated on dual treatment of hydroxychloroquine and mycophenolate mofetil and is currently recovering. Discussion We relooked at this immunology when he was first diagnosed with primary APS in 2017. He was strongly positive for ANA at titre of 1:640 but did not have manifestations of SLE then. We researched literature on the predictors for primary APS to develop into SLE. Both disease spectrums have several overlapping manifestations including haemolytic anaemia, thrombocytopenia, lymphopenia, neurological symptoms, renal manifestations and livedo reticularis. Freire et al (1) retrospectively analysed 80 patients with primary APS and found 14 (17.5%) progressed to develop APS-SLE in 5.2 +/- 4 years. ANA positivity was found in 100% those who eventually developed APS-SLE. Other specific autoantibodies including anti-dsDNA, anti-ribosomal P, anti-Ro, anti-La and anti-U1RNP are found more commonly in the cohort too. In their APS-SLE cohort, they also found a correlation with younger age at the time of diagnosis of APS and the duration of the disease. Andreoli L et al (2) demonstrated anti-nucleosome antibody (anti-NCS) was frequently found in their cohort of APS-SLE patients but no relationship between clinical-serological was found, except for a weak correlation between anti-dsDNA antibodies. 2 of their primary APS who developed severe SLE had displayed high titres of anti-NCS years before. Tarr T et al (3) found 4 of their primary APS who developed SLE all presented with cerebrovascular events as the thrombotic manifestation. Key learning points • Primary APS has been associated with the development of SLE • Certain antibody profile such as ANA, anti-nucleosome antibody may predict the development of SLE in primary APS patients • Younger age, cerebrovascular events as thrombotic manifestation at diagnosis of primary APS and the length of duration of primary APS may be associated with the development of SLE at a later stage

An immuno-inflammatory profiling of asymptomatic individuals in a malaria endemic area in Uganda

Malaria caused by Plasmodium falciparum leads to the destruction of red blood cells (RBCs). A better understanding of how naturally immune individuals control infections should be valuable for future vaccine studies. Antibodies against RBCs and RBC surface antigens were measured together with different inflammatory markers in healthy adults living in a malaria endemic area of Uganda and compared to Swedish healthy adults. Antibodies binding to RBCs were clearly elevated in Ugandans compared to Swedish samples, and for RBC surface antigens the Ugandans had higher levels of antibodies against JMH, but not against Cromer or Kell. Twenty-eight percent of the Ugandans were PCR-positive for P. falciparum, and these had higher levels of IgG against parasite extract and more inhibition in functional growth/invasion assays, but levels of antibodies against RBC, RBC surface antigens, results from Direct Antiglobulin Tests (DAT) and indirect antiglobulin tests were similar when compared with PCR-negative individuals. When inflammatory markers (α-1-antitrypsin, haptoglobin, orosomucoid/α-1-acid glycoprotein, CRP, IgG, IgA and IgM) were measured there were in general almost no signs of inflammation except for clearly elevated levels of IgG. Some had low levels of haptoglobin and for orosomucoid more than half of the individuals had clearly reduced levels. There was no correlation between the inflammatory markers and PCR-positivity, antibodies against RBCs or parasites. In conclusion, for healthy adults living in a malaria endemic area, there was a clear presence of antibodies against RBCs in parallel with high levels of IgG and almost no signs of inflammation, even though many individuals were carrying parasites.