1. RBC Methodologies-IIBOND KING

2. I.Erythrocyte sedimentation rate(ESR) Rate of settling of RBC from the plasma after the additionof anticoagulant.Importance of ESR1. Good index for the presence of hidden carcinoma butactive diseases.2. It measures the suspension stability of RBC.3. It measures the abnormal concentration of fibrinogenand serum globulin.Roleaux formation (Packing or piling of RBC) 3. MethodsA.Wintrobe and landsberg method Anticoagulant used- Ammonium potassium oxalate (wintrobe solution/ double oxalate/balanced oxalate/ paul-Hellers soln.) Tube wintrobe tube- Left side red; 0 on top and 10 cm bottom.- Right side white; 10 cm top and 0 bottom. 4. Procedure1. With a long stem pasteur pipet, fill the wintrobe tube with oxalated blood up to 0 mark.2. Let the wintrobe tube stand perfectly vertical.3. Read result after 1 hour. Reading must be done on the left red side of the tube.Normal values Male - (0-9) mm/hr Female - ( 0-20 )mm/hr ( bcg less RBC ) Children- (1-13) mm/hr 5. Red White 010 Layers -Plasma layer -Buffy coat (WBC and platelets) -Packed RBC (hematocrit)Wintrobetube Red cells at bottom 0 at bottom10 6. Females have more space in settle down and faster thanmale and children because they have less RBC. ( 1cm 10mm/hr)B.Westergren method (200mm) most sensitive andmost accurate . - Anticoagulant used -3.8% sodium citrate - Tube- westergren tube (through suction method longtube) 7. 200 mm0 8. Procedure1. Fill the tube with the citrated blood2. Stand the tube vertically and read result at the end of the 1st hrs. and 2nd hrs.Normal values Male (3-5) mm/hr 7-15 mm/2hr Female (4-7) mm/hr12-17 mm/2hr 9. ComparisionWintrobe Wester grenBore3 mm 2.5 mmGraduationup to 100 mm up to 200 mmAnticoagulant Double oxalate3.8% sodium citrateAmount of 1 ml2.4 mlbloodReading once TwiceHematocrit 10. C. Graphic cutler Anticoagulant 3.8% sodium citrateD. Linzenmeier Anticoagulant 3.8% sodium citrateE. Roarke- Ernstiene Anitocagulant HeparinF. Brays Anticoagulant- 3.8% sodium citrate 11. G. Micro methods1) Micro landau Anticoagulant- 5% sodium citrate2) Smith micro Anticoagulant- 5% sodium citrate3) Crista or hellige- vollmerStages of ESR1. Initial rouleaux formation (first 10 min)2. Period of rapid settling (next 40 min)3. Period of final settling (last 10 min) total 60 minutes or 1hr. 12. Factors in ESR1. Intrinsic Factor- nos of RBC ( less RBC faster settlement)- size of RBC ( Bigger the size is faster the settlement)- viscosity of Plasma ( less viscous fast settlement)* nos of RBC- inversely* size of RBC- directly 13. 2. Extrinsic factor Length of tube ( smaller length fast settlement) Diameter of tube (wider diameter fast settlement) Position of tube(vertical or slightly fast settlement Temperature ( high temp. fast settlement) Pipetting ( incorrect pipetting result error) Volume of blood ( less blood faster settle.) Anticoagulant (more anticoagulant slow settlement) 14. II Osmotic fragility test Test the stability of RBC in hypotonic solutions. Follows the law of osmosis. Factor affecting OFT - Red cell shape - Red cell volume - Red cell surface Area - State of Red cell membrane *Fragile cells( decrease)- spherocytes *Resistant cells( increase)- sickle cell , target cell,reticulocytes 15. Methods1.Sanford method Different conc. Of hypotonic solution 12 test-tube is used Initial solution used 0.5% NaclInterpretation No hemolysis tubes with compact sediment and clearsolution. Initial hemolysis -1st tube from the left with not socompact sediment and with dark red solution Complete hemolysis - 1st tube from the left withoutsediment and with dark red solution. 16. Normal values Initial hemolysis- tube 22 Complete hemolysis- tube17Increase OFT Initial hemolysis- tube 24 ( increased-hemolytic anemia ,hereditary spherocyte) Complete tube 20 { decrease-sickle cell anemia,thalassemia , jaundice, SIDA(severe ion deficiencyAnemia)}.Decreased OFT Initial hemolysis-tube 19 Complete hemolysis-tube 15 17. 2. Modified Sanford in terms of ml 18. 3. Griffin and Sanford method4. Dacies method Hemolysis read is used through spectrophotometer. (Transparent fake pink- light pink- red) 19. III.Erythrocyte Indices Important in assessing borderline types of anemia. Computed using 3 determinants Hb, hematocrit and RBCcount.A. Mean corpuscular volume (MCV) Average volume of an individual RBC.volume % Hct x 10 = cubic micra or femtoliterRBC in millions Normal value- 82- 92 cubic micra. 20. Interpretation 95- 160 cubic micra- macrocyte 72-79 cubic micra microcyte 50-71 cubic micra microcyte hypochromic(less Hb)Example Hct = 46 vol % RBC count 5,000,000/ cumm MCV= 46 x 10 = 92 cubic micra5 21. B. Mean Corpuscular Hemoglobin (MCH) Ratio of Hb to red cell count Average weight or amount of Hb in an individual RBC inmillions gm Hb x 10 = uug or picogram RBC in million Normal value (27-33) uugInterpretation> 33 uug- macrocyte< 27uug microcyte< 22 uug microcytic hypochromicExampleHb = 16gm/100mlRBC count = 5,500,000/cummMCH= 16 x 10 = 29 uug 5.5 22. C. Mean corpuscular hemoglobin conc.(MCHC) Mean conc. Of Hb in the average RBC.Normal Value = 32-38% Average weight or amount of Hb in an individual RBC gm Hb x 100 = % vol. % HctExample Hb -16 gm/ 100ml Hct = 46 vol. % MCHC = 16 x 1000 = 34.7%46 23. Normal normochromic < 32 hypochromic > 38 hyperchromic