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Immunohematology and Genetic Testing (red cells, leukocytes and platelets)

(P-IG-41) Two Novel JK Alleles Discovered by Investigation of Discrepant Serology and Genotyping Results

Sunday, October 20, 2024
12:00 PM - 1:00 PM  
Background/Case Studies: Discrepancies between test methods, such as serology and genotyping, have led to identification of novel alleles. More than thirty alleles that code for altered or silenced expression of Jk antigens are known to-date. In this study, we investigated samples from a donor and a patient that typed Jk(a−) by serology and were predicted Jk(a+) by targeted genotyping.

Study

Design/Methods: Phenotyping was performed by standard tube methods with monoclonal Anti-Jka and -Jkb. Genomic DNA was isolated from WBCs in whole blood (Qiagen QIAmp Mini Kit). PreciseType HEA (Werfen/Immucor) and Sanger sequencing of JK (exons 3 to 10 and flanking intron regions) were performed. 

Results/Findings:

Sample 1 was from a female Caucasian donor. RBCs typed Jk(a−b+) with Ortho BioClone Anti-Jka (MS15) and Anti-Jkb (MS8), but PreciseType HEA predicted Jk(a+b+). Sequencing revealed that the sample was heterozygous for c.838G >A (p.Asp280Asn), indicative of JK*01/02, silent variant c.588A >G (p.Pro196=), c.582C >G (p.Tyr194Ter), which defines null allele JK*01N.03, and novel silent variant c.582C >T (p.Tyr194=), listed on gnomAD v.4.1.1 (rs34756616) with an overall frequency of 0.00001847. 

Sample 2 was from a pregnant female, ethnicity unknown. RBCs typed Jk(a−) with Ortho BioClone Anti-Jka (MS15), but PreciseType HEA predicted Jk(a+). Additional phenotyping was performed to resolve the discrepancy; the sample was found to be Jk(a−) with Immucor and Bio-Rad Anti-Jka. Sequencing revealed that the sample was heterozygous for c.838G >A, indicative of JK*01/02, silent variant c.588A >G (p.Pro196=), and novel variant c.1043T >A, which leads to a premature stop codon (p.Leu348Ter) and is listed on gnomAD v.4.1.1 (rs2047661090) with an overall frequency of 0.000006570. Fresh EDTA sample was not available for adsorption/elution studies

Conclusions: In sample 1 we identified a rare and peculiar genotype consisting of two different variants at the same coding position, on defining null allele JK*01N.03, the other novel allele JK*02(582T). In sample 2 we identified novel possibly-null allele JK*01(1043A). The c.1043A variant encodes a stop codon, consistent with a null phenotype. These cases highlight the limitations of targeted genotyping, emphasize the importance of resolving serology/DNA discrepancies, and demonstrate the complementarity of serology and genomics for accurate blood typing.