November 18, 2025

HLA-B27 is a genetic marker associated with immune conditions including ankylosing spondylitis and reactive arthritis. Detection of HLA-B27 can be accomplished through flow cytometric evaluation of phenotype (using lymphocytes in a fresh blood sample), or by PCR analysis of genomic DNA (also taken from leukocytes in a peripheral blood sample). Historically, Warde has performed HLA-B27 flow cytometry, with a sendout test available for situations requiring the genetic assay instead. PCR testing had a higher cost to the client and a longer turn-around time. Clinical performance for the two methods is comparable in most cases; exceptions are discussed below.

We are happy to announce that Warde has developed an in-house PCR assay that will be available at similar cost to the flow cytometry HLA-B27 test. The required specimen type is whole blood in EDTA tubes, with stability up to 7 days refrigerated – a significant improvement over flow cytometry. As such, for most patients we suggest PCR as the more robust option with less risk of sample cancellation due to delay in transport. The new test code is B27M; we recommend communicating this change to providers at your institution who frequently order test code B27 (HLA-B27 by flow cytometry).

For the most prevalent HLA-B27 alleles, and of course for patients without HLA-B27, performance characteristics for flow cytometry and PCR are similar. PCR tends to have better overall sensitivity and specificity. The PCR assay detects the following specific HLA-B*27 family alleles according to primer alignments:

B*27:01	B*27:02:01-03	B*27:04:01, :02, :04	B*27:05:02-28	B*27:06	B*27:08-10
B*27:12-13	B*27:15-18	B*27:23	B*27:25-29	B*27:31	B*27:35
B*27:37-42	B*27:44-58	B*27:59N	B*27:60-63	B*27:64N	B*27:65N
B*27:66N	B*27:67-69	B*27:71-80	B*27:82-89	B*27:91-93	B*27:94N
B*27:95	B*27:96:01-02	B*27:97-101	B*27:103-124

The following uncommon HLA-B*27 alleles are expected to be missed according to alignments:

B*27:04:03	B*27:07:01-04	B*27:11	B*27:14	B*27:19-21
B*27:24	B*27:30	B*27:32-34	B*27:36	B*27:43
B*27:70	B*27:81	B*27:90:01-02	B*27:102	B*27:125

Based on information from the reagent manufacturer, the flow cytometric assay shows the potential for cross-reactivity with HLA-B7, resulting in some false positives when the HLA-B7 antigen is present and HLA-B27 is absent. Additionally, many laboratories using the same flow cytometry reagent as Warde show false negative results for HLA-B27 type 27:03, a phenomenon we have observed here as well.

In summary, options available for HLA-B27, as of November 18, include:

HLA-B27 genotyping by PCR –  test code: B27M

HLA-B27 flow cytometry (more stringent stability requirements) – test code: B27

B27M is recommended.