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Biomedical subjects

K Sparks

Publications and source records attributed to K Sparks.

At least 37 records · Page 2Linked to original sources
PubMed1988

Immunochemical analysis of acetaminophen covalent binding to proteins. Partial characterization of the major acetaminophen-binding liver proteins.

A sensitive immunoassay for detecting acetaminophen (APAP) bound to proteins was developed using an affinity purified antibody directed against the N-acetylated end of the APAP molecule. Western blots of electrophoretically resolved liver proteins taken from mice given an hepatotoxic dose of APAP demonstrated that nearly 85% of the total detectable protein-bound APAP was covalently associated with proteins of 44 and 58 kD. Pretreatment of liver extracts with the sulfhydryl-specific reagent, N-ethylmaleimide (NEM), prior to derivatization with the reactive metabolite of APAP, N-acetyl-p-benzoquinone imine (NAPQI), greatly reduced immunochemically detectable APAP-protein adducts and indicated that the antibody detects protein-thiol conjugates of APAP. To investigate the basis of the binding selectivity in vivo, a variety of systems which yielded APAP-protein adducts were analyzed. Systems which activate APAP enzymatically, as in hepatocyte suspensions or in post-mitochondrial (S9) fractions fortified with an NADPH-regenerating system, resulted in a protein binding profile similar to that produced in vivo. Conversely, when extracts or cells were treated with chemically synthesized NAPQI, an alternative protein binding profile was obtained. Two-dimensional electrophoretic analysis of the reduced protein thiol (PSH) content of liver proteins using [3H]NEM labeling revealed that the 58 kD APAP-binding proteins were rich in PSH, whereas the major 44 kD binding protein had virtually no detectable PSH. Many PSH-rich proteins that were not arylated in vivo did bind NAPQI in vitro. However, the 44 kD proteins were not arylated when chemically synthesized NAPQI was added to homogenates or cell suspensions. The present data further suggest that, in addition to the amount and reactivity of free protein sulfhydryls, the cellular localization with respect to the cytochrome P-450 activation site may influence the susceptibility of proteins to NAPQI binding. These findings signal the need for caution in interpreting studies of APAP mechanisms that rely solely on NAPQI addition.

Acetaminophen↗
PubMed1983

IgG specific antibodies to rye grass and ragweed pollen antigens in the tear secretions of patients with vernal conjunctivitis.

We studied the tears of 30 patients (25 males and five females, ranging in age from 8 to 34 years) with vernal conjunctivitis for pollen-specific IgG antibodies to rye grass and ragweed antigen E by an enzyme-linked immunosorbent assay (ELISA). Eighteen of 30 (60%) and 20 of 30 (67%) patients with vernal conjunctivitis had significant levels (more than 2 S.D. from the mean of control tears) of IgG antibodies in their tear secretions to rye grass (geometric mean = 68.7 ELISA units) and ragweed antigen E (geometric mean = 50 ELISA units), respectively. In contrast, the control groups (eight atopic individuals, four with seasonal rhinitis, and 12 with allergic conjunctivitis) had low amounts of specific IgG antibodies to these two pollen antigens in their tears. Total IgG and IgM were also increased in the tears of patients with vernal conjunctivitis. To evaluate whether these immunoglobulins and specific IgG antibodies were locally produced by the conjunctival tissues, we used transferrin as a marker for the leakage of plasma proteins into tears. We found that the specific IgG antibodies to rye grass or ragweed antigen E, or both, in the tears were locally produced by the conjunctival tissues. The local production ranged from 20% to 99.9%. Of 17 patients with vernal conjunctivitis and undetectable pollen-specific IgE antibodies in their tears, 14 (82.4%) had tear specific IgG antibodies to rye grass or ragweed antigen E or both, whereas of those with measurable tear IgE antibodies, only nine of 13 (69%) had tear IgG antibodies to rye grass or antigen E or both. These results suggested that both IgE- and IgG-mediated immune mechanisms may be important in the pathogenesis of vernal conjunctivitis.

Adolescent↗
PubMed1983

Rapid quantitative microenzyme-linked immunosorbent assay for tetanus antibodies.

A rapid quantitative microenzyme-linked immunosorbent assay for the detection of immunoglobulin G antibodies to tetanus toxoid is described. The assay is performed in 2.5 h and detects antibody levels of between 0.1 and 6 U/ml. Quantitation of the antibody concentration is unique in that multiple serum dilutions are tested on a single patient, the results are plotted as an antibody dilution curve, and the area under the curve is integrated for a final concentration. Tests for immune status on sera from 77 individuals demonstrated tetanus antibody concentrations of greater than or equal to 0.1 U/ml in 92% of a pediatric group and 62% of an adult group. The microenzyme-linked immunosorbent assay procedure is reproducible and is useful for the determination of immune status and immunization response.

Adult↗