Expedient screening for HIV-1 protease inhibitors using a simplified immunochromatographic assay.
A colloidal gold-based immunochromatographic (IC) strip test was developed and validated for the detection of HIV-1 protease (HIV-PR) activity and inhibitory effect of HIV-PR inhibitors (PIs). It is a unique 'two-step' process requiring the combination of proteolysis of HIV-PR and an immunochromatographic reaction. Monoclonal antibodies to the free C-terminus of HIV matrix protein (HIV-MA) conjugated to gold particles and a monoclonal antibody against intact and cleaved forms of the HIV-MA are immobilized on the 'Test'-line of the IC strip. Using lopinavir, a potent HIV protease inhibitor, the IC-strip was optimized to detect inhibitory activity against HIV-protease. At a lopinavir concentration of 1000ng/mL (its suggested minimum effective concentration), a HIV-PRH6 concentration of 6mg/mL and incubation period of 60min were the optimal conditions. A preliminary comparison between a validated high-performance liquid chromatography assay and the IC-strip to semi-quantify HIV protease inhibitor concentrations (lopinavir and atazanavir) demonstrated good agreement. This simplified method is suitable for the rapid screening of novel protease inhibitors for future therapeutic use. Moreover, the IC strip could also be optimized to semi-quantify PIs concentrations in plasma samples.
Center for Research and Innovation, Faculty of Medical Technology, Mahidol University, Bangkok 10700, Thailand.
Kuntida
K
Kitidee
Division of Clinical Immunology, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand.
Wannisa
W
Khamaikawin
Division of Clinical Immunology, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand; i+MED Laboratories Company Limited, Bangkok, Thailand.
Weeraya
W
Thongkum
Program for HIV Prevention and Treatment (PHPT/IRD 174), Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand.
Yardpiroon
Y
Tawon
Program for HIV Prevention and Treatment (PHPT/IRD 174), Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand; Harvard T.H Chan School of Public Health, Boston, MA 02115, USA.
Tim R
TR
Cressey
Researcher in Pharmaceutics, R&D Institute, Government Pharmaceutical Organization, Bangkok 10400, Thailand.
Rachaneekorn
R
Jevprasesphant
Division of Clinical Immunology, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand; Biomedical Technology Research Unit, National Center for Genetic Engineering and Biotechnology, National Science and Technology Development Agency at the Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand.
Watchara
W
Kasinrerk
Division of Clinical Immunology, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand; Center of Biomolecular Therapy and Diagnostic, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai 50200, Thailand. Electronic address: asimi002@hotmail.com.
Chatchai
C
Tayapiwatana
D000818:Animals; D055910:Antibodies, Immobilized / Q000378:metabolism*; D000911:Antibodies, Monoclonal / Q000378:metabolism*; D002846:Chromatography, Affinity / Q000379:methods*; D005260:Female; D017979:Gold Colloid / Q000737:chemistry; D016333:HIV Protease; D017320:HIV Protease Inhibitors / Q000097:blood* / Q000378:metabolism; D006801:Humans; D061466:Lopinavir / Q000097:blood; D051379:Mice; D008807:Mice, Inbred BALB C
D055910:Antibodies, Immobilized; D000911:Antibodies, Monoclonal; D017979:Gold Colloid; D017320:HIV Protease Inhibitors; D061466:Lopinavir; D016333:HIV Protease; C514803:p16 protease, Human immunodeficiency virus 1
HIV-1 protease inhibitor; Immunochromatographic strip; Rapid screening platform; Semi-quantification