Title:Development of an Isothermal Point-of-care Genetic Rapid Test for
the Detection of the HLA-B*57:01 Allele, a Predictor for Hypersensitivity
Reaction Caused by Abacavir, for Stratifying Patients for Antiretroviral
Abacavir HIV Therapy
Volume: 20
Issue: 1
Author(s): Alexander E. Jacobsen*, Antje Jüngling, Corinna Gorges and Moritz Eidens
Affiliation:
- Department of Sales & Marketing, Mainz Biomed Germany GmbH, Mainz, Germany
Keywords:
Abacavir, HIV therapy, hypersensitivity reaction, HLA-B*57:01, isothermal recombinase-polymeraseamplification, isothermal RPA, lateral flow dipstick, lateral flow test.
Abstract:
Background: Abacavir is used in the treatment of HIV-infected patients. A hypersensitive
reaction (HSR) occurs in about 5-8% of patients treated with Abacavir. The
HLA-B*57:01 allele is a valuable predictor for HSR and its screening is mandatory prior to
treatment with Abacavir.
Objective: Current screening methods require considerable investments for equipment. In
order to lower the required investments and enable physician practices to perform the
screening in a point-of-care (PoC) setting, our objective was to develop a novel isothermal
genetic rapid test that requires a minimal setup cost, does not require specific training and
thus is suitable for a physician practice setting.
Methods: We developed an isothermal recombinase polymerase amplification (RPA) for the
specific amplification of the HLA-B*57:01 allele using allele-specific primers coupled to Biotin.
Primers specific for human lactase gene, coupled to Digoxigenin, were used as an internal
amplification control (IAC). Lateral flow dipstick provided rapid and accurate detection of
HLA-B*57:01 allele and IAC via the respective antibodies sprayed on the strips surface.
Results: The reference method identified the HLA-B*57:01 allele in the reference sample,
in 2 out of 28 buccal swab samples and in 2 out of 13 blood samples. The initial isothermal
RPA resulted in unspecific amplification of the HLA-B*57:01 allele. By further optimization
steps the specific amplification of the allele and the detection on lateral flow dipstick
was observed. The newly developed isothermal RPA was validated.
Conclusion: The method developed fulfils the requirements for a genetically based PoC
screening system for the HLA-B*57:01 variant, requiring a minimal investment for a heating
block and a pipette.
