Biomedical subjects

G E Valkirs

Publications and source records attributed to G E Valkirs.

4 recordsLinked to original sources

PubMed1986

Internally referenced ImmunoConcentration assays.

We have developed a method for incorporating an internal reference zone in addition to the test zone in a single immunoConcentration device. We have illustrated the method by constructing an internally referenced assay for human choriogonadotropin (HCG) in serum. The internally referenced assay, which can be performed in about 5 min, provides a measured overall precision (CV) of approximately 15% at a concentration, in serum, of approximately 50 int. units/L. The test zone consists of an anti-HCG antibody adsorbed onto latex microspheres and entrapped within a porous support. The reference zone contains antibody against alkaline phosphatase (EC 3.1.3.1), which has also been adsorbed onto latex microspheres and trapped within the support. The use of an internal reference moderates the effects of variations in assay conditions on the interpretation of assay results.

Alkaline Phosphatase↗

PubMed1985

ImmunoConcentration--a new format for solid-phase immunoassays.

A new format for solid-phase immunoassays has been developed in which a monoclonal antibody-coated membrane, incorporated into a cylindrical, disposable device, regulates sample and reagent delivery. We illustrate the method with a two-site, immunoenzymometric assay that can detect human choriogonadotropin at less than 50 int. units/L (4 micrograms/L) in urine and less than 25 int. units/L (2 micrograms/L) in serum and takes less than 5 min to perform. The solid-phase antibody is located in a circular area in the center of the membrane so that in the presence of the hormone, after addition of substrate, a blue enzyme product is generated in this circular area. The high ratio of surface area to volume within the microporous matrix of the membrane assures short diffusion distances and therefore rapid binding of liquid-phase reagents to the solid phase. Pseudo-first-order reaction kinetics describe the binding of antigen to immobilized antibody and the binding of enzyme-labeled antibody to immobilized antigen. The speed and simplicity of this format may facilitate testing for many analytes, both soluble and particulate, as well as serological testing.

Alkaline Phosphatase↗

PubMed1983

Interaction of cytochrome c with reaction centers of Rhodopseudomonas sphaeroides R-26: localization of the binding site by chemical cross-linking and immunochemical studies.

The location of the cytochrome binding site on the reaction center of Rhodopseudomonas sphaeroides was studied by two different approaches. In one, cross-linking agents, principally dithiobis(propionimidate) and dimethyl suberimidate, were used to link cytochrome c and cytochrome c2 to reaction centers; in the other, the inhibition of electron transfer by antibodies against the subunits was investigated. Cytochrome c (horse) cross-linked to the L and M subunits, whereas cytochrome c2 (R. sphaeroides) cross-linked only to the L subunit. The cross-linked reaction center-cytochrome complexes were isolated by affinity chromatography. The rate of electron transfer in the cross-linked cytochrome c2 complex was the same as that in the un-cross-linked complex. However, when cytochrome c was used, the rate in the cross-linked complex was about 15 times slower than that in the un-cross-linked complex. Fab fragments of antibodies specific against the L and M subunits blocked electron transfer from both cytochrome c (horse) and cytochrome c2 (R. sphaeroides). Antibodies specific for the H subunit did not block either reaction. We conclude that the cytochrome binding site on the reaction center is close (approximately 10 A) to both the L and M subunits, possibly in a cleft between them.

Binding Sites↗

PubMed1982

Topography of reaction center subunits in the membrane of the photosynthetic bacterium, rhodopseudomonas sphaeroides.

The localization of the reaction center polypeptides (L, M, and H) in the membranes of both the wild-type, strain 2.4.1, and the carotenoidless mutant, R-26, of Rhodopseudomonas sphaeroides was determined by using affinity-purified antibodies specific for these proteins. Binding of the antibodies to reaction center subunits in spheroplasts was visualized in the electron microscope by immunoferritin labeling. The H and M subunits were labeled at both the cytoplasmic and the periplasmic surfaces of the membrane, whereas the L subunit was labeled only at the periplasmic surface of the membrane. Thus, the reaction center is asymmetrically oriented in the membrane with at least two subunits (H and M) spanning the membrane.

Antibodies, Bacterial↗