Biomedical subjects

R A Fredrickson

Publications and source records attributed to R A Fredrickson.

6 recordsLinked to original sources

PubMed1999

Improved detection of bovine herpesvirus 1 in artificially infected bovine semen by protein amplification.

Infection with bovine herpesvirus 1 (BHV 1) occurs worldwide and causes serious economic losses due to loss of animals, abortions, decreased milk production, and loss of body weight. There is a real need for sensitive diagnostic procedures for detection of the presence of virus in order to achieve effective control of BHV 1-induced diseases. BHV 1 is frequently found in bovine semen and can be widely transmitted through artificial insemination. Thus the detection of BHV 1 in artificial insemination centers and semen banks is of crucial importance in the control of its dissemination to the cattle industry, worldwide. In the present study, a protein amplification assay following polymerase chain reaction (PCR) of the highly conserved BHV 1 glycoprotein D gene was used in order to improve the sensitivity of direct virus detection in bovine semen. This method of BHV 1 detection is at least 200 orders of magnitude more sensitive than traditional PCR and would have direct clinical applications in antigen-based detection tests. In this method, amplification of the BHV 1 gD gene by PCR is followed by a coupled in vitro transcription translation of a small aliquot from the reaction. When the transcription translation was carried out in the presence of [35S]methionine and the products analyzed by SDS PAGE and autoradiography, 0.0014 TCID50 of virus could be detected in raw bovine semen in contrast to 0.28 TCID50 of virus detected using traditional PCR. Given the limitations in the method used for protein detection, this 'in vitro protein amplification' has the potential of attaining superior sensitivity for direct virus detection in clinical samples.

Animals↗

PubMed1999

Characterization of monoclonal antibodies against bovine herpesvirus 1 gD fusion protein expressed in E. coli.

A total of 20 hybridoma cell lines secreting monoclonal antibodies (MAbs) against E. coli expressed bovine herpesvirus-1 (BHV-1) gD fusion protein were produced following the fusion of Sp2/0 myeloma cells with splenocytes from BALB/c mice immunized previously with immunoaffinity purified BHV-1 gD fusion protein. An indirect fluorescent antibody test (IFAT) using BHV-1 infected MDBK cells was used for the selection of positive hybridomas secreting specific antibody. The monoclonal antibody isotypes were 11 IgM, six IgG2b, one IgG1 and two IgG3. All MAbs reacted positively with the E. coli expressed BHV-1 gD fusion protein, BHV-1 infected MDBK cell lysates and PCR BHV-1 gD transcription-translation polypeptide antigens by an ELISA.

Animals↗

PubMed1999

Purification and partial characterization of a cadmium-binding protein from the liver of rainbow trout (Onchorynchus mykiss).

This study describes the isolation and partial characterization of a low molecular weight (approximately 14 kDa), cadmium-binding protein from rainbow trout (Onchorynchus mykiss) liver. Rainbow trout were injected intraperitoneally with 3.5 mg/kg cadmium chloride (total body dose) twice weekly for 3 wk. Livers were removed and a cadmium-binding protein was isolated. Monoclonal antibodies produced against this protein were used in the affinity purification process. Amino acid analysis showed the protein contained 3.8 mol% cysteine, 3.5 mol% phenylalanine, 2.2 mol% tyrosine and 1.9 mol% histidine. The low cysteine content suggests that it was distinct from metallothionein. The monoclonal antibodies were also used to identify the protein in liver homogenates from both cadmium-exposed and control fish and in the testes of cadmium-exposed mice lacking the gene for both metallothionein-1 and metallothionein-II. The compound identified in this study represents a non-metallothionein cadmium-binding protein that appears to be highly conserved.

Animals↗

PubMed1990

A unitized enzyme-labeled immunometric digoxin assay suitable for rapid testing.

An enzyme-labeled immunometric assay has been developed for measuring digoxin concentrations in serum or plasma. Unitized, compartmentalized reagents are used with an automated sample-processing instrument. The enzyme activity of the processed sample, which is directly proportional to the digoxin concentration, is measured by using a reagent strip and the Ames Seralyzer reflectance photometer. The test takes less than 15 min, and digoxin concentrations are calculated from a two-point calibration line stored in the instrument. Within-run CVs for controls at four concentrations ranged from 2.3% to 3.8%; between-run CVs were from 1.5% to 2.6%. Results obtained with clinical serum samples correlated well (r greater than 0.96) with those obtained by fluorescent polarization immunoassay (Abbott TDx) and RIA (Clinical Assays and NML). This rapid and convenient method for monitoring digoxin concentrations in serum or plasma is particularly well suited for decentralized sites such as emergency rooms, urgent-care centers, and physicians' offices.

Antibodies, Monoclonal↗

PubMed1989

Improved procedure for preparation of F(ab')2 fragments of mouse IgGs by papain digestion.

Preparation of F(ab')2 fragments from mouse monoclonal IgG by papain digestion can result in incorporation of traces of papain into antibody fragments by disulfide exchange. Such trace contamination can have detrimental effects on the integrity of these antibody fragments following reduction. Gel filtration and ion exchange chromatography procedures did not eliminate papain contamination from F(ab')2 preparations. The use of antibody specific for papain to remove this contamination from F(ab')2 preparations is described.

Animals↗

PubMed1984

Peptide mapping of bovine pancreatic ribonuclease A by reverse-phase high-performance liquid chromatography. I. Application to the reduced and S-carboxymethylated protein.

Complete peptide maps of reduced and S-carboxymethylated ribonuclease A were obtained by reverse-phase high-performance liquid chromatography with the following peptide-chain cleavage techniques: cyanogen bromide cleavage, limited and extensive Staphylococcus aureus protease digestion, tryptic digestion, and tryptic followed by chymotryptic digestion. Commercial samples of S. aureus protease exhibited a broader specificity than had previously been reported, as demonstrated by its ability to cleave after glutamine residues. Cleavage after asparagine and serine residues was also strongly implicated. The procedures developed require roughly 0.1 to 1 mg of ribonuclease A for the peptide mapping of this protein. These procedures will be useful for the identification of the sites of a chemical modification and also for the isolation of a variety of peptides for further studies.

Amino Acid Sequence↗