Biomedical subjects

C F Pincombe

Publications and source records attributed to C F Pincombe.

4 recordsLinked to original sources

PubMed1986

Competitive solid-phase enzyme immunoassay for measuring digoxin in serum.

In this clinically useful enzyme immunoassay of digoxin in serum, we mix sample, beta-galactosidase-labeled digoxin, and anti-digoxin Fab fragments for 30 min at room temperature, then use Sepharose-bound second antibody for phase separation, and measure the unbound enzyme activity directly in the supernate of the equilibrium reaction mixture. The immunoassay buffer--phosphate-buffered isotonic saline with added rabbit globulin (4 g/L), hydrolyzed gelatin (2 g/L), Brij 96 detergent (5 g/L), glycerol (0.25 mol/L), and N-acetyl-8-anilinonaphthalene-1-sulfonic acid (2 mmol/L)--minimizes serum matrix effects for convenient measuring of unbound enzyme--digoxin conjugate. The immunoassay developed with Fab fragments has better displacement characteristics than that with intact antibody. Performance of the assay compares favorably with that of other manual digoxin immunoassays; in comparison studies with EMIT involving 110 clinical specimens, the coefficient of correlation was 0.97.

Antibody Formation↗

PubMed1985

Ligand displacement immunoassay--demonstration of its use for the measurement of serum phenobarbital and phenytoin.

We describe a clinically useful ligand displacement immunoassay for the measurement of serum or plasma phenobarbital and phenytoin. A conjugate of drug-specific antiserum covalently bound to micro-crystalline cellulose and a beta-galactosidase-labelled drug derivative is prepared as a lyophilized reagent. Sample is added to the conjugate and, after 5-min incubation at room temperature, the bound and displaced enzyme-ligand are separated by brief centrifugation. The enzyme activity of the displaced enzyme-ligand is measured on a spectrophotometer or centrifugal analyser. Linear calibration curves are obtained with appropriate sample dilution, allowing generation of the standard curve with a saline zero reference and a single calibrator. The performance of the assay compared favourably with other immunoassays; comparison studies with clinical specimens gave a correlation with EMIT of 0.96 (n = 122) for phenobarbital and 0.95 (n = 96) for phenytoin.

Humans↗

PubMed1985

The use of a monosaccharide linkage group in a heterologous solid-phase enzyme immunoassay for phenytoin.

The preparation is described of monosaccharide-hapten derivatives containing a galactosyl linkage group which bears little structural resemblance to the straight-chain hydrocarbons commonly used for attaching haptens to proteins for use as immunogens. These derivatives are readily synthesised and are coupled to enzymes under mild conditions to produce bridge-heterologous enzyme immunoassays. The use of beta-galactosidase-monosaccharide-phenytoin derivatives in the development of sensitive phenytoin enzyme immunoassays is described and the assays are compared with hapten-heterologous, site-heterologous and homologous phenytoin enzyme immunoassays prepared using other phenytoin derivatives. This technique has application in the development of immunoassays for haptens which have only one functional group to which linkage groups can be attached for covalent coupling of the hapten to proteins.

Animals↗

PubMed1984

Ligand displacement immunoassay: a novel enzyme immunoassay demonstrated for measuring theophylline in serum.

We describe a ligand displacement immunoassay for measurement of theophylline in serum or plasma and show it to be clinically useful. A conjugate of theophylline-specific antiserum covalently bound to micro-crystalline cellulose and a beta-galactosidase-labeled theophylline derivative is prepared as a lyophilized reagent. Sample is added to the conjugate and, after 5-min incubation at room temperature, the bound and displaced enzyme-ligand are separated by brief centrifugation. The enzyme activity of the displaced enzyme-ligand is measured on a spectrophotometer or centrifugal analyzer. Linear calibration curves are obtained with appropriate sample dilution, allowing generation of the standard curve with a saline zero reference and a single calibrator. Performance of the assay compares favorably with that of other theophylline assays; comparison studies with use of 145 clinical specimens gave a coefficient of correlation with EMIT of 0.97.

Anticoagulants↗