Weak D and DEL alleles detected by routine SNaPshot genotyping: identification of four novel RHD alleles

Transfusion. 2011 Feb;51(2):401-11. doi: 10.1111/j.1537-2995.2010.02830.x. Epub 2010 Aug 16.


Background: Molecular RHD blood group typing is very efficient for managing donors and patients carrying any of the various molecular types of weak D and DEL. The purpose of the work was to develop a multiplex polymerase chain reaction (PCR) SNaPshot assay for simultaneous detection of weak D and DEL alleles that are prevalent in Europeans, Africans, and Asians.

Study design and methods: Preliminary profiling was carried out on single-nucleotide polymorphisms (SNPs) associated with 13 prevalent RHD alleles, that is, weak D Types 1, 2, 3, 4.0, 4.0.1, 4.1, 4.2, 5, 11, 15, and 17; RHD(IVS3+1g>a); and RHD(K409K). Multiplex PCR was used to amplify six RHD regions encompassing 14 SNPs. Identification was obtained by incorporation of the complementary dye single base at the 3'-end of each probe-primer. A prospective analysis was then carried out on 152 blood samples from patients (n = 53) and donors (n = 88) with equivocal RhD serology and pregnant women (n = 11).

Results: After validation, our SNaPshot assay allowed direct genotyping of 82.9% of samples overall and 100% of samples harboring weak D Types 1, 2, 3, and 4.1 alleles. In the remaining 17.1% of samples overall, sequence investigation allowed accurate genotyping. In addition, four novel RHD alleles were identified, that is, RHD(S256P), RHD(L390L), RHD(F410V), and RHD(IVS4-2a>g).

Conclusion: The SNaPshot assay described herein is a helpful supplementary tool for resolving doubtful RhD serology. By allowing accurate identification of weak D and DEL alleles this assay should allow better management of the donors and the patients genotyped weak D Types 1, 2, 3, and 4.1 who can receive D+ blood units.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Agglutination Tests
  • Alleles*
  • Amino Acid Substitution
  • Blood Donors
  • Blood Grouping and Crossmatching / methods*
  • Consensus Sequence / genetics
  • Exons / genetics
  • Female
  • France
  • Genotype
  • Humans
  • Polymerase Chain Reaction / methods*
  • Polymorphism, Single Nucleotide*
  • Pregnancy
  • Prospective Studies
  • RNA Splice Sites / genetics
  • Rh-Hr Blood-Group System / chemistry
  • Rh-Hr Blood-Group System / genetics*


  • RNA Splice Sites
  • Rh-Hr Blood-Group System
  • Rho(D) antigen