Development and evaluation of an immunoassay for the monitoring of the anti-HIV drug amprenavir.
An assay for routine therapeutic drug monitoring of anti-HIV HAART drugs in clinical use is highly desirable, in order to rapidly measure the pharmacokinetic parameters on single patients. We have started a project to develop a panel of enzyme-linked immunosorbent assays (ELISA) for the whole set of HAART drugs, and the development, performance and evaluation of the assay for amprenavir is described here. A diazo conjugate of amprenavir has been used in order to raise polyclonal anti-amprenavir antibodies in rabbits. Antisera have been used to set up a quantitative and rapid competitive assay. Plasma samples are simply diluted in the assay buffer after thermal inactivation, before running the assay. The assay allows the detection of amprenavir in the quantification range 400-5000 ng/ml, in a diluted plasma sample. The assay has been compared with an HPLC reference technique, on 27 samples from treated patients. Within the quantification range, the ELISA data are well correlated with the HPLC results by a regression line close to the identity, and a Bland-Altman analysis shows the agreement between the two methods.
Bastiani
Erica
E
Biostrands s.r.l., via del Follatoio 12, I-34148 Trieste, Italy. e.bastiani@biostrands.com
Benedetti
Fabio
F
Berti
Federico
F
Campaner
Pietro
P
Donadel
Elena
E
Montagna
Michela
M
Regazzi
Mario
M
Rinaldi
Serena
S
Savoini
Adriano
A
Venturini
Romina
R
D000328:Adult; D000818:Animals; D019380:Anti-HIV Agents / Q000097:blood* / Q000737:chemistry / Q000627:therapeutic use; D000906:Antibodies / Q000097:blood / Q000276:immunology; D002219:Carbamates / Q000032:analysis / Q000097:blood* / Q000276:immunology / Q000627:therapeutic use; D002851:Chromatography, High Pressure Liquid; D003429:Cross Reactions / Q000276:immunology; D016903:Drug Monitoring / Q000379:methods*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005663:Furans; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D015394:Molecular Structure; D010755:Organophosphates / Q000032:analysis / Q000276:immunology; D010047:Ovalbumin / Q000737:chemistry / Q000276:immunology; D011817:Rabbits; D015203:Reproducibility of Results; D012710:Serum Albumin, Bovine / Q000276:immunology; D013449:Sulfonamides / Q000032:analysis / Q000097:blood* / Q000276:immunology / Q000627:therapeutic use; D014611:Vaccination
D019380:Anti-HIV Agents; D000906:Antibodies; D002219:Carbamates; D005663:Furans; D007106:Immune Sera; D010755:Organophosphates; D013449:Sulfonamides; D012710:Serum Albumin, Bovine; C095108:amprenavir; D010047:Ovalbumin; C426859:fosamprenavir