Related Experiment Videos
Expression of alpha-tryptase and beta-tryptase by human basophils
Sherryline Jogie-Brahim1, Hae-Ki Min, Yoshihiro Fukuoka
1Division of Rheumatology, Allergy and Immunology, Department of Internal Medicine, Virginia Commonwealth University, PO Box 980263, Richmond, VA 23298, USA.
Background:
Alpha and beta-tryptase levels in serum are clinical tools for the evaluation of systemic anaphylaxis and systemic mastocytosis. Basophils and mast cells are known to produce these proteins.
Objective:
The current study examines the effect of the alpha,beta-tryptase genotype on basophil tryptase levels and the type of tryptase stored in these cells.
Methods:
Tryptase extracted from purified peripheral blood basophils from 20 subjects was examined by using ELISAs measuring mature and total tryptase and by using an enzymatic assay with tosyl-Gly-Pro-Lys-p-nitroanilide. Tryptase genotypes (4:0, 3:1, and 2:2 beta/alpha ratios) were assessed by using a hot-stop PCR technique with alpha,beta-tryptase-specific primers. Total alpha,beta-tryptase mRNA was measured by means of competitive RT-PCR, and ratios of alpha to beta-tryptase mRNA were measured by means of hot-stop RT-PCR.
Results:
Tryptase in all but one of the basophil preparations was mature and enzymatically active. Tryptase quantities in basophils were less than 1% of those in tissue mast cells. Tryptase genotypes (beta/alpha) among the 20 donors were 4:0 in 7, 3:1 in 7, and 2:2 in 6. Tryptase protein and mRNA levels per basophil were not affected by the tryptase genotype.
Conclusion:
Basophils from healthy subjects contain modest amounts of mature and enzymatically active tryptase unaffected by the tryptase genotype.