Flow injection chemiluminescent immunoassay with para-phenylphenol and sodium tetraphenylborate

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Analytica Chimica Acta485(2003)

57–62

Flow injection chemiluminescent immunoassay with

para-phenylphenol and sodium tetraphenylborate

as synergistic enhancers

Jian-Xin Luo1,Xiu-Cen Yang∗

West China School of Pharmacy,Sichuan University,Chengdu610044,PR China

Received18November2002;received in revised form12March2003;accepted24March2003

Abstract

A new approach ofﬂow injection chemiluminescent immunoassay(FICLIA)with synergistic enhancement was developed. The synergistic action was signiﬁcant in the chemiluminescence(CL)system of luminol–H2O2–HRP with two enhancers, para-phenylphenol(PPP)and sodium tetraphenylborate(NaTPB).The relative CL intensity was128.84%stronger(compared to NaTP

B alone)and122.66%stronger(compared to PPP alone)using both enhancers than any one alone(both P<0.001). While the present approach was applied to the determination of rabbit IgG as a model analyte,satisfactory results were obtained in the dynamic range of2–60␮g l−1with a detection limit of0.68fmol per injection and a precision of4.7–9.3%, and the recoveries were92.5–99.4%.The results of rabbit IgG determination accorded with those of radio immunoassay (RIA).The lifetime of the immuno-column was considerably long:one column was applied to more than200determinations in1month without signiﬁcant loss of activity.

Keywords:Flow injection analysis;Chemiluminescent analysis;Immunoassay;Synergistic action;para-Phenylphenol;Sodium tetraphenylborate

1.Introduction

For a long period of time,radio immunoassay (RIA)has been the most sensitive assay,which was ﬁrst introduced in1959[1].However,strong efforts have been undertaken in regard to the replacement of radioactive substances in immunoassays by other de-tection approaches.The enzyme immunoassay(EIA)∗Corresponding author.

E-mail address:xcyang@(X.-C.Yang).

1Present address:Chengdu Medical College,The Third Military Medical University,Chengdu610083,PR China.was introduced in1971[2,3],which,in terms of sen-sitivity,was for theﬁrst time a match for the RIA. In the EIA,several enzymes,alkaline phosphatase (AP),␤-d-galactosidase and horseradish peroxidase (HRP)were often used as labels.The detection of these labels can be carried out in various ways, including photometric,ﬂuorimetric or chemilumino-metric techniques after enzymatic conversion of a suitable substrate into a detectable product.Despite the success of the enzyme-linked immunosorbent assay(ELISA),its end use is limited due to the re-quirement of trained persons and long assay time[4]. These limitations have sparked the combination of