Prevalence of Anti- A and Anti – B Haemolysins among blood group ‘O’ donors in Makurdi, Nigeria.
Description
Blood group O donors are “inappropriately” called universal donors. These donors could become potentially “dangerous” if haemolysins are detected in their plasma. This study determined the prevalence of anti–A and anti–B haemolysins among blood group O donors in Makurdi, Benue State. Three hundred and five voluntary group O donors were screened for anti–A and anti–B haemolysins using the standard tube technique and samples showing haemolysis were titrated for anti A and anti B haemolysins. The overall prevalence of anti–A and/or anti–B haemolysins was 66.2%. Prevalence of anti–A haemolysins was 6%, anti–B haemolysins 14%, and both was 45.6% of blood donors.
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5mls of serum was obtained from clotted samples and screened for anti–A and anti–B haemolysins using the standard tube technique . The haemolytic properties of IgG anti–A and anti–B was adopted for this test. Fresh A and B cells were washed four times in saline and 5% red cell suspension was made. Three tubes were arranged, and two parts of washed A, B and O cells were dispensed into each of the three tubes. The O cells were used as negative control. Four parts each of fresh group O serum (initially stored at –20oC and thawed) were added to the tubes and then incubated at 370C for one hour. Subsequently, the tubes were centrifuged at 2500rpm for a minute and then held before a source of light. With minimal disturbance, the supernatant was observed for haemolysis macroscopically, and the results were recorded. Haemolysis was graded as follows: complete haemolysis 3+, partial haemolysis 2+, trace haemolysis 1+, and no visual haemolysis, negative. In samples that showed haemolysis, anti-A and anti-B haemolysins were titrated thus: 300µl of donor serum (neat) was placed in the first well of the first row of a 12 by 8 microplate well. Next, 150µl of normal saline was placed in subsequent wells (after the neat) to the 11th well. Thereafter, 150 µl of serum from the neat well was then transferred to the second well, thoroughly mixed to give 1 in 2 dilution of the haemolysin. 150µl of the mixture in the second well was transferred to the third well, thoroughly mixed, to give 1 in 4 dilution of haemolysin. The procedure was repeated for the subsequent wells to give 1 in 8 up to 1 in 1024 dilutions of the haemolysin. The procedure described above for the 1st row was also carried out for the second and third rows. To the wells of the first row was added 5% saline suspension of A cells. To the second and third row wells were added 5% saline suspension of B and O cells respectively. O cells served as negative control and the well containing it (row 3 wells) served as serum blank well. All microplates were incubated at 37°C for an hour. The supernatant of each well was visually examined for haemolysis with the visual titre taken as the last serum dilution where haemolysis was seen. For wells where there was no observable visual haemolysis, spectrophotometric evidence of haemolysis was demonstrated. Following visual assessment of haemolysis, other microplates were set up corresponding with the ones with no visible haemolysis. 120 µl of saline was placed into each well. 30 µl microlitres of the supernatant from the corresponding incubated wells above was pipetted into each of the new wells and then thoroughly mixed to produce a 1 in 5 dilution of each suspension (corresponding to 1 in 200 dilution of the red cell suspension). The optical density of each suspension was read on a spectrophotometer at 540 nm using saline as blank. The spectrophotometric titre was taken as the dilution just before that which gave the same optical density as the serum blank.