Dried blood spots of pooled samples for RHD gene screening in blood donors of mixed ancestry.
OBJECTIVE
In this study, we present a strategy for RHD gene screening based on real-time polymerase chain reaction (PCR) using dried blood spots of pooled samples.


BACKGROUND
Molecular analysis of blood donors may be used to detect RHD variants among the presumed D-negative individuals. RHD genotyping using pooled samples is a strategy to test a large number of samples at a more reasonable cost.


METHODS
RHD gene detection based on real-time PCR using dried blood spots of pooled samples was standardised and used to evaluate 1550 Brazilian blood donors phenotyped as RhD-negative. Positive results were re-evaluated by retesting single samples using real-time PCR and conventional multiplex PCR to amplify five RHD-specific exons. PCR-sequence-specific primers was used to amplify RHDψ allele.


RESULTS
We devised a strategy for RHD gene screening using dried blood spots of five pooled samples. Among 1550 serologically D-negative blood donors, 58 (3.74%) had the RHD gene. The non-functional RHDψ allele was detected in 47 samples (3.02%).


CONCLUSIONS
The present method is a promising strategy to detect the RHD gene among presumed RhD-negative blood donors, particularly for populations with African ancestry.
Fundação Centro de Hematologia e Hemoterapia de Minas Gerais, Hemominas, Minas Gerais, Brazil.
M C F
MC
Silva-Malta
Fundação Centro de Hematologia e Hemoterapia de Minas Gerais, Hemominas, Minas Gerais, Brazil.
N C Fidélis
NC
Araujo
Fundação Centro de Hematologia e Hemoterapia de Minas Gerais, Hemominas, Minas Gerais, Brazil.
O V Neves
OV
Vieira
Fundação Centro de Hematologia e Hemoterapia de Minas Gerais, Hemominas, Minas Gerais, Brazil.
L Cayres
LC
Schmidt
Fundação Centro de Hematologia e Hemoterapia de Minas Gerais, Hemominas, Minas Gerais, Brazil.
P de Cassia
Pde C
Gonçalves
Fundação Centro de Hematologia e Hemoterapia de Minas Gerais, Hemominas, Minas Gerais, Brazil.
M Lobato
ML
Martins
D001782:Blood Donors; D001788:Blood Grouping and Crossmatching / Q000379:methods*; D046148:Donor Selection / Q000379:methods*; D059788:Dried Blood Spot Testing / Q000379:methods*; D005260:Female; D006801:Humans; D008297:Male; D012204:Rh-Hr Blood-Group System / Q000235:genetics*
D012204:Rh-Hr Blood-Group System; C026410:Rho(D) antigen
Rh blood group; blood donors; immunohematology; molecular testing