Detection of HIV-1 p24 based on gold nanoparticle probe and PCR
10.3760/cma.j.issn.1009-9158.2012.05.013
- VernacularTitle:金纳米探针结合PCR检测HIV-1 p24抗原的研究
- Author:
Huahuang DONG
;
Jianli LIU
;
Hong ZHU
;
Guiyun ZHANG
;
Lingzhang MENG
;
Wenge XING
;
Maofeng QIU
;
Yao XIAO
;
Jun YAO
;
Pinliang PAN
;
Yan JIANG
- Publication Type:Journal Article
- Keywords:
HIV-1 core protein p24;
Gold nanoparticles;
Polymerase chain reaction;
Electrophoresis,agar gel
- From:
Chinese Journal of Laboratory Medicine
2012;35(5):448-452
- CountryChina
- Language:Chinese
-
Abstract:
ObjectiveTo establish a novel assay for HIV-1 p24 ultrasensitive detection based on Gold Nanoparticle Probe (GNP) and PCR.MethodsSandwich ELISA method was established by a pair of anti-p24 monoclonal antibodies (mAbs),1G12 and 1D4,and was used to detect recombinant HIV-1 p24 antigen.The bio-barcode DNA was 47 bp,selected from genome of Arabidopsis,and formed double-stranded DNA by hybridization with the capture DNA (complementary with bio-barcode DNA) modified with sulfhydryl.Then double-stranded DNA were conjugated on the surface of 1D4-modified gold nanoparticles by sulfhydryl,and the Gold Nanoparticle Probe was produced.1G12 was precoated in the micropaltes,and in the presence of target recombinant HIV-1 p24 protein,a sandwich immuno-complex would form by adding GNP.Then the bio-barcode DNA in the immuno-complex were released by heating as detection signal,and consequently characterized by the polymerase chain reaction (PCR) with synthesized special primers and analyzed by 4％ agar gel electrophoresis,so HIV-1 p24 antigen could be evaluated.The sensitivity comparison between the new assay and ELISA can be done.ResultsSandwich ELISA was used to quantify HIV-1 p24 antigen by monoclonal antibodies 1G12 and 1D4,and the limit of detection (LOD) was 1000 pg/ml.The new GNP assay was established by the same pair of antibodies,combined with PCR and agar gel electrophoresis,and was used to indirectly detect HIV-1 p24 antigen.The band intensity of PCR products paralleled with the quantity of HIV-1 p24 antigen,and the limit of detection (LOD) could reach down to 1 pg/ml.ConclusionThe new assay based on GNP and PCR was efficient in the detection of HIV-1 p24,which is at least 3 orders of magnitude more sensitive than traditional ELISA.
