<prompt> A 23-year-old female presented with jaundice and pallor for 2 months. Her peripheral blood smear shows the presence of spherocytes. The most relevant investigation to arrive at a diagnosis is;Reticulocyte countOsmotic fragility testCoombs testTests fro PNH  <response>: Ans. c. Coombs test Ref: Robbins 9e p643-644; Chandrasoma and Taylor 3e p401History of jaundice in a 23-year old female with pallor and spherocytes in peripheral blood points towards the diagnosis of autoimmune hemolytic anemia AIHA. The diagnostic test for AIHA is the antiglobulin test Coombs test.Autoimmune hemolytic anemia AIHA:Onset is very often abrupt and can be dramatic.The hemoglobin level can drop, within days, to as low as 4gdL; the massive red cell removal will produce jaundice, and often the spleen will be enlarged. When this triad is present, the suspicion of AIHA must be high.The diagnostic test for AIHA is the antiglobulin test Coombs test.AIHA is diagnosed by detection of autoantibodies with the direct antiglobulin direct Coombs test.IgG is present in warm antibody hemolytic anemia, and C3 C3b and C3d in cold antibody disease.Reticulocyte count is increased in both hereditary spherocytosis and AIHA.Osmotic Fragility Test is used to evaluate hemolytic anemia, especially hereditary spherocytosis, and immune hemolytic states.Test for PNH:PNH will present with hemoglobinuria.Blood tests in PNH show changes consistent with intravascular hemolytic anemia:Low hemoglobinRaised LDHRaised reticulocytes immature red cells released by the bone marrow to replace the destroyed cellsRaised bilirubin a breakdown product of hemoglobinDecreased levels of haptoglobinThe direct antiglobulin test DAT, or direct Coombs test is negative, as antibodies do not cause the hemolysis in PNH.Warm antibody hemolytic anemiaCold antibody hemolytic anemia Antibodies here bind with antigens on RBC at body temperature 37deg C These antibodies are called warm antibodies and are nearly IgGQ Antibodies here bind with antigens on RBC better at temperature lower than 37degC These antibodies are called reactive antibodies and are usually IgM Rarely IgG antibodies e.g. The Donath Landsteiner antibody of Paroxysmal cold hemoglobinuriaQCausesCauses1. Idiopathic2. Lymphomas: CLLQ, Non-Hodgkins etc.3. SLEQ and other Collagen Vascular DiseasesQ4. Drugs: Alpha-MethyldopaQ Penicillin type stable pattern Quinidine type unstable pattern1. Acute: Mycoplasma InfectionQ, Infections mononucleosisQ2. Chronic: IdiopathicQ, Lymphoma3. Paroxysmal cold hemoglobinuriaQ IgGMechanism of HemolysisMechanism of Hemolysis Human red cells cooled with IgG are trapped by splenic macrophages leading to red cell destructionQ Antibodies of IgM type bind on red cell surface and cause agglutination Hemolytic effect is mediated through fixation of C3 to RBC surfaceQ.DiagnosisDiagnosis Positive direct Coombs test, at 37degC for presence of warm antibodies on surface of Red cell. Positive Indirect Coombs test at 37degC for presence of large quantities of warm antibodies in serum.Q Positive direct Coombs test for detection of C3 on the red cell surface, but IgM responsible for coating on red cells is not found.Q 