Hydroquinone diphosphate: an alkaline phosphatase substrate that does not produce electrode fouling in electrochemical immunoassays.
Hydroquinone diphosphate (HQDP) was synthesized and compared to phenyl phosphate (PP) and 1-naphthyl phosphate (NP) as a substrate for alkaline phosphatase (AP) under electrochemical immunoassay (EIA) conditions. Voltammetric and amperometric experiments showed that electrochemical oxidation of hydroquinone (HQ), which is the AP hydrolysis product of HQDP, did not produce electrode passivation, even with repeated biosensor use. In contrast, phenol and 1-naphthol, the hydrolysis products of PP and NP, respectively, were shown to be irreversibly oxidized on the electrode surfaces, and produced rapid electrode passivation, resulting in complete loss of electrode signal. When employed as AP substrate in an iridium oxide based EIA, HQDP produced significantly larger amperometric responses (117 microA/cm2) compared to PP (31 microA/cm2) and NP (27 microA/cm2). The results presented in this paper show that HQDP is an attractive alternative to commonly used AP substrates such as NP and PP. The substrate shows excellent hydrolytic stability, produces larger amperometric responses (than PP or NP), and does not produce sensor passivation.
EIC Laboratories, Inc., 111 Downey Street, Norwood, MA 02062, USA. mwilson@eiclabs.com
Michael S
MS
Wilson
R David
RD
Rauh
D000469:Alkaline Phosphatase / Q000032:analysis* / Q000737:chemistry*; D004563:Electrochemistry / Q000295:instrumentation* / Q000379:methods; D004566:Electrodes; D004789:Enzyme Activation; D004868:Equipment Failure; D019544:Equipment Failure Analysis; D006868:Hydrolysis; D006873:Hydroquinones / Q000032:analysis* / Q000737:chemistry*; D007118:Immunoassay / Q000295:instrumentation* / Q000379:methods; D007074:Immunoglobulin G / Q000032:analysis*; D010755:Organophosphates / Q000032:analysis* / Q000737:chemistry*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013379:Substrate Specificity
D006873:Hydroquinones; D007074:Immunoglobulin G; D010755:Organophosphates; C105856:hydroquinone 1,4-bisphosphate; D000469:Alkaline Phosphatase