Vagner Castro

Investigation of red blood cell antigens with highly fluorescent and stable semiconductor quantum dots

We report a new methodology for red blood cell antigen expression determination by a simple labeling procedure employing luminescent semiconductor quantum dots. Highly luminescent and stable core shell cadmium sulfide/cadmium hydroxide colloidal particles are obtained, with a predominant size of 9 nm. The core-shell quantum dots are functionalized with glutaraldehyde and conjugated to a monoclonal anti-A antibody to target antigen-A in red blood cell membranes. Erythrocyte samples of blood groups A+, A2+, and O+ are used for this purpose. Confocal microscopy images show that after 30 min of conjugation time, type A+ and A2+ erythrocytes present bright emission, whereas the O+ group cells show no emission. Fluorescence intensity maps show different antigen expressions for the distinct erythrocyte types. The results obtained strongly suggest that this simple labeling procedure may be employed as an efficient tool to investigate quantitatively the distribution and expression of antigens in red blood cell membranes.

A prospective study on the prevalence and risk factors for neonatal thrombocytopenia and platelet alloimmunization among 9332 unselected Brazilian newborns

BACKGROUND: Neonatal thrombocytopenia (NT) occurs in 0.5 to 0.9% of unselected Caucasian newborns. However, the prevalence of this complication in other populations is unknown. In this study the prevalence/causes of NT was determined in Brazilian newborns, a population characterized by admixture among Indigenous, Africans, and Caucasians.

The new noninvasive occlusion spectroscopy hemoglobin measurement method: a reliable and easy anemia screening test for blood donors

BACKGROUND: The tests used for anemia screening in blood donors are based on fingerstick samples, leading to discomfort and complaints. The aim of this study was to analyze the feasibility of occlusion spectroscopy method in blood banks and to compare the method with fingerstick hemoglobinometer and hemoglobin (Hb) determination on an automatic blood analyzer.

Association of the immature platelet fraction with sepsis diagnosis and severity.

Management of Sepsis would greatly benefit from the incorporation of simple and informative new biomarkers in clinical practice. Ideally, a sepsis biomarker should segregate infected from non-infected patients, provide information about prognosis and organ-specific damage, and be accessible to most healthcare services. The immature platelet fraction (IPF) and immature reticulocyte fraction (IRF) are new analytical parameters of the complete blood count, that have been studied as biomarkers of several inflammatory conditions. Recently, a study performed in critically-ill patients suggested that IPF could be a more accurate sepsis biomarker than C-reactive protein (CRP) and procalcitonin. In this retrospective study we evaluated the performance of IPF and IRF as biomarkers of sepsis diagnosis and severity. 41 patients admitted to two intensive care units were evaluated, 12 of which with severe sepsis or septic shock, and 11 with non-complicated sepsis. Significantly higher IPF levels were observed in patients with severe sepsis/septic shock. IPF correlated with sepsis severity scores and presented the highest diagnostic accuracy for the presence of sepsis of all studied clinical and laboratory parameters. No significant differences were observed in IRF levels. Our results suggest that IPF levels could be used as a biomarker of sepsis diagnosis and severity.

Identification of platelet refractoriness in oncohematologic patients

OBJECTIVES: To identify the occurrence and the causes of platelet refractoriness in oncohematologic patients. INTRODUCTION: Platelet refractoriness (unsatisfactory post-transfusion platelet increment) is a severe problem that impairs the treatment of oncohematologic patients and is not routinely investigated in most Brazilian services. METHODS: Forty-four episodes of platelet concentrate transfusion were evaluated in 16 patients according to the following parameters: corrected count increment, clinical conditions and detection of anti-platelet antibodies by the platelet immunofluorescence test (PIFT) and panel reactive antibodies against human leukocyte antigen class I (PRA-HLA). RESULTS: Of the 16 patients evaluated (median age: 53 years), nine (56%) were women, seven of them with a history of pregnancy. An unsatisfactory increment was observed in 43% of the transfusion events, being more frequent in transfusions of random platelet concentrates (54%). Platelet refractoriness was confirmed in three patients (19%), who presented immunologic and non-immunologic causes. Alloantibodies were identified in eight patients (50%) by the PIFT and in three (19%) by the PRA-HLA. Among alloimmunized patients, nine (64%) had a history of transfusion, and three as a result of pregnancy (43%). Of the former, two were refractory (29%). No significant differences were observed, probably as a result of the small sample size. CONCLUSION: The high rate of unsatisfactory platelet increment, refractoriness and alloimmunization observed support the need to set up protocols for the investigation of this complication in all chronically transfused patients, a fundamental requirement for the guarantee of adequate management.

A novel association of acquired ADAMTS13 inhibitor and acute dengue virus infection.

BACKGROUND: Dengue is a mosquito‐borne viral disease with an increasing incidence worldwide. Thrombocytopenia is a common finding in dengue virus (DV) infection; however, the underlying mechanisms remain unknown.

Severe immune haemolysis in a group A recipient of a group O red blood cell unit

Summary.  Haemolysis caused by passive ABO antibodies is a rare transfusional complication. We report a case of severe haemolytic reaction in a 38‐year‐old man (blood group A) with lymphoma who had received one red blood cell (RBC) unit group O. After transfusion of 270 mL, the patient experienced fever, dyspnoea, chills and back pain. On the following morning he was icteric and pale. Haptoglobin was inferior to 5.8 mg dL−1, haemoglobin was not increased and lactate dehydrogenase was elevated. Haemolysis was evident on observation of the patients post‐transfusion samples. The recipients red cells developed a positive direct antiglobulin test and Lui elution showed anti‐A coated the cells. Fresh donor serum had an anti‐A titre of 1024, which was not reduced by treating the serum with dithiothreitol. Donor isoagglutinin screening has been determined by microplate automated analyser and showed titre higher than 100. Physicians should be aware of the risk of haemolysis associated with ABO‐passive antibodies. There is generally no agreement justifying the isoagglutinin investigation prior to transfusion. However, automated quantitative isoagglutinin determination could be part of the modern donor testing process, mainly in blood banks where identical ABO group units (platelets or phenotyped RBCs) are not available owing to limited supply.

Platelet antibody detection by flow cytometry: an effective method to evaluate and give transfusional support in platelet refractoriness

Background Immune platelet refractoriness is mainly caused by human leukocyte antigen antibodies (80-90% of cases) and, to a lesser extent, by human platelet antigen antibodies. Refractoriness can be diagnosed by laboratory tests and patients should receive compatible platelet transfusions. A fast, effective and low cost antibody-screening method which detects platelet human leukocyte/platelet antigen antibodies is essential in the management of immune platelet refractoriness. Objective The aim of this study was to evaluate the efficiency of the flow cytometry platelet immunofluorescence test to screen for immune platelet refractoriness. Methods A group of prospective hematologic patients with clinically suspected platelet refractoriness treated in a referral center in Campinas, SP during July 2006 and July 2011 was enrolled in this study. Platelet antibodies were screened using the flow cytometry platelet immunofluorescence test. Anti-human leukocyte antigen antibodies were detected by commercially available methods. The sensitivity, specificity and predictive values of the immunofluorescence test were determined taking into account that the majority of antiplatelet antibodies presented human leukocyte antigen specificity. Results Seventy-six samples from 32 female and 38 male patients with a median age of 43.5 years (range: 5-84 years) were analyzed. The sensitivity of the test was 86.11% and specificity 75.00% with a positive predictive value of 75.61% and a negative predictive value of 85.71%. The accuracy of the method was 80.26%. Conclusion This study shows that the flow cytometry platelet immunofluorescence test has a high correlation with the anti-human leukocyte antigen antibodies. Despite a few limitations, the method seems to be efficient, fast and feasible as the initial screening for platelet antibody detection and a useful tool to crossmatch platelets for the transfusional support of patients with immune platelet refractoriness.

Platelet associated IgG may be related with thrombocytopenia in patients with myelodysplastic syndromes.

Thrombocytopenia is common in patients with myelodysplastic syndromes (MDS) and immune destruction of platelets could be an important factor for its occurrence. We prospectively analyzed platelet-associated IgG (PAIgG) through platelet immunofluorescence test (PIFT), mean platelet volume (MPV), platelet size deviation width (PDW) and glycocalicin index (GCI) of 54 patients with MDS, classified according to the International Prognostic Scoring System (IPSS). Thrombocytopenia (platelet count<100×10(9)/L) was correlated with a higher amount of PAIgG, significantly higher MPV and increased GCI. In addition, worse prognosis IPSS groups were associated with a higher positivity of PIFT, which could be indicative of advanced disease.

Diagnosis of Scott syndrome in patient with bleeding disorder of unknown cause.

Scott syndrome is a rare bleeding disorder due to an impaired exposure of phosphatidilserine on the platelet membrane, compromising the platelet procoagulant activity, thrombin generation and, thus, the clot formation. We report a case of a 17-year-old female adolescent with bleeding episodes of unknown cause. She had normal coagulation, but altered platelet aggregation under arteriolar flow, indicating platelet dysfunction. Furthermore, the expression of Annexin V was markedly reduced and the diagnosis of Scott syndrome was established. She was treated with platelet transfusions and demonstrated a clinical improvement. Scott syndrome may be investigated in cases with bleeding history and normal coagulation tests.