Determination of hemoglobin Bart's in alpha thalassemia traits by two-site immunoradiometric assay. I. Purification of hemoglobin Bart's and preparation of specific anti-Hb Bart's.
This research report describes methods for the preparation of hemolysate and the isolation and purification of hemoglobins Bart's, A, A2, E, F and H. Procedures for the preparation of anti-Hb Bart's by injecting purified Hb Bart's into rabbits is indicated in the time schedule. The rabbit antisera were evaluated by antigen-antibody reaction in agar gel. Although the antiserum reacted with Hb Bart's but not with Hb A, A2,E and H, it also cross-reacted with Hb F. After the rabbit antisera were absorbed with Hb F, the antisera were highly specific because it only reacted with Hb Bart's. The purified specific anti-Hb Bart's was labelled with radioactive 125I by chloramine-T method. After passing through Sephadex G-100 column, the 125I labelled specific anti-Hb Bart's was obtained in the first peak. This radioactive labelled anti-Hb Bart's was ready to use in the two-site immunoradiometric assay.
Makonkawkeyoon
L
L
Department of Biochemistry, Faculty of Medicine, Chiang Mai University, Thailand.
Ongchai
S
S
Sanguansermsri
T
T
D004588:Electrophoresis, Cellulose Acetate; D000939:Epitopes; D006455:Hemoglobins, Abnormal / Q000276:immunology / Q000302:isolation & purification*; D006801:Humans; D005779:Immunodiffusion; D007122:Immunoelectrophoresis; D015592:Immunoradiometric Assay; D007457:Iodine Radioisotopes; D017085:alpha-Thalassemia / Q000235:genetics*
D000939:Epitopes; D006455:Hemoglobins, Abnormal; D007457:Iodine Radioisotopes; C030448:hemoglobin Bart's