A method was developed for the purification of kallikrein from human urine. The procedure consisted of three steps: ultradialysis, diethyl-(2-hydroxypropyl) aminoethyl (QAE) ion exchange radial flow membrane chromatography and affinity chromatography on aprotinin agarose. It is simple and suitable for large-scale purification. The purified product was checked by SDS-PAGE and matrix-assisted laser desorption:ionization mass spectrometery (MALDI). A single band with apparent molecular weight (MW) 42,000 in SDS-PAGE and a single monomer peak with MW33,000 in MALDI were observed, respectively. The biological activity tested by ELISA showed positive immunological identity of the purified product compared with human urinary kallikrein standard.

Amino Acid Sequence↗

PubMedDate not supplied

Characterization of monoclonal and polyclonal antibodies to human tissue kallikrein.

Monoclonal antibodies to purified human urinary kallikrein have been developed. Selection of antibody producing clones was based on 125I-kallikrein binding activity of hybridoma media in both radioimmunoassay and enzyme-linked immunosorbent assay. Three clones (2 IgG1, 1 IgG2b) were subcloned, characterized, and compared with the polyclonal antiserum generated in rabbits immunized with the purified kallikrein. With radioimmunoassay, mouse ascitic fluids or rabbit antisera dilutions showing 50% binding to 125I-kallikrein were 1:1.2 X 10(6) (E7A9), 1:1.2 X 10(5) (H6A6), 1:8.0 X 10(4) (E12H1), and 1:1.4 X 10(6) (the rabbit antisera). With enzyme-linked immunosorbent assay, mouse ascitic fluids from clones E7A9 and H6A6 showed half-maximal absorbance at dilutions of 1:2.1 X 10(5) and 1:1.0 X 10(5) respectively, and the polyclonal antiserum showed half-maximal absorbance at a dilution of 1:2.0 X 10(4). These monoclonal antibodies showed no cross-reactivity with rat tissue kallikrein, rat urinary plasminogen activator, or dog pancreatic kallikrein, while the polyclonal antiserum showed some cross-reactivity. The binding of monoclonal or polyclonal antibodies to 125I-human urinary kallikrein was not affected by human plasma kallikrein, thrombin, or urokinase in a competitive radioimmunoassay. By using purified human urinary kallikrein immobilized to agarose, antibodies produced by clones E7A9 and H6A6 and in the rabbit antisera were purified to homogeneity. Each of these affinity-purified antibodies inhibited the esterase activity, and two of the three inhibited the kininogenase activity, of human urinary kallikrein. A sandwich immunosorbent assay was developed to measure this kallikrein using monoclonal antibody from the clone E7A9 in conjunction with the polyclonal antibodies.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗

PubMedDate not supplied

Diffuse annular fusiform adenocarcinoma in a dog.

Canine colonic intestinal adenocarcinoma typically presents as rectal polypoid or annular stenotic masses causing clinical signs consistent with large bowel disease. This report discusses an unusual case of intestinal adenocarcinoma in an 11-year-old, neutered male German shepherd dog presented for evaluation of anorexia, profuse watery diarrhea, and weight loss. In this dog, colonic adenocarcinoma diffusely infiltrated the entire large bowel and caused an annular fusiform lesion, as confirmed by endoscopic biopsies and postmortem examination. Other unique features included a paucity of desmoplasia associated with the neoplastic lesion and widespread metastasis to regional lymph nodes, lung, and prostate.

Adenocarcinoma↗

PubMedDate not supplied

Continuity of care: incorporating patient perceptions.

Continuity of medical care has been conceptualized by some researchers as an attitude on the part of the patient and provider rather than as a succession of visits to the same provider. A perception of continuity (PC) scale containing 23 statements was developed to empirically test this conceptualization. A cross-sectional, random sample survey of adult patients in an established suburban family practice utilized a chart review and mailed patient questionnaire to assess the potential value of this scale. The PC scale had a high degree of internal reliability and a greater correlation with patient satisfaction than two commonly used provider continuity measures, even after controlling for demographic variables. The cost of ambulatory primary medical care was not significantly related to any of the continuity measures. The measurement of patient attitudes to define continuity provided information distinct from provider continuity formulas.

Adolescent↗