The Development of a Radioimmunoassay Procedure for the Estimation of Tamm—Horsfall Glycoprotein in Human Serum

1977 ◽  
Vol 52 (2) ◽  
pp. 183-191 ◽  
Author(s):  
P. J. G. Avis
Keyword(s):  
Human Serum ◽  
Serum Proteins ◽  
Solid Phase ◽  
Normal Serum ◽  
Small Sample ◽  
Agarose Beads ◽  
Assay Procedure ◽  
Series Of Experiments ◽  
Sepharose 4B ◽  
Cyanogen Bromide Activation

1. A quantitative radioimmunoassay was developed for the measurement of nanogram amounts of Tamm—Horsfall (TH) glycoprotein in the presence of serum proteins at concentrations above 30 mg/ml. 2. Specific anti-(TH-glycoprotein) antibodies were labelled with 125I and these were usable for a period of 8 weeks. 3. Agarose beads (Sepharose 4B), to which TH-glycoprotein had been coupled via cyanogen bromide activation of the Sepharose, were used as the solid phase in the assay. This proved stable upon storage at 4°C for periods in excess of 4 months. 4. The dissociation of the glycoprotein in the presence of serum proteins that was necessary for quantification was achieved by subjecting the sample to ultrasonication. 5. Assays conducted on a small sample of normal serum produced evidence that normal serum contained amounts (50–180 ng/ml) of a substance that reacted similarly to TH-glycoprotein in the assay procedure and in a series of experiments conducted to confirm the presence of this substance in human serum.

scholarly journals Capacity of human serum to depolymerize actin filaments

Blood ◽  
1987 ◽  
Vol 70 (2) ◽  
pp. 524-530
Author(s):  
PA Janmey ◽  
SE Lind
Keyword(s):  
Human Serum ◽  
Actin Filaments ◽  
Serum Proteins ◽  
Normal Serum ◽  
Slow Phase ◽  
Rapid Phase ◽  
Plasma Gelsolin ◽  
Preferential Formation

Human blood depolymerizes filamentous (F-)actin. The interaction of actin filaments and monomers with human serum was studied by following the kinetics and extent of the depolymerization of pyrene-labeled F- actin and by analysis of serum proteins adhering to immobilized actin monomers. In physiologic Ca2+ concentrations, the depolymerization of F- actin proceeds in two stages: a rapid phase, attributed to direct severing of filaments by plasma gelsolin, and a slow phase attributed to the binding of actin monomers to vitamin D-binding protein (DBP). Without Ca2+, only the slow phase is observed. Human serum can completely depolymerize 10 to 18 mumol/L of actin, of which approximately 5 mumol/L occurs rapidly. Depolymerization can be accounted for by the normal serum concentrations of gelsolin and DBP. Fibrin(ogen) and fibronectin, which bind actin in vitro, do not contribute to the kinetics or extent of its depolymerization. Affinity chromatography and functional assays for the presence of gelsolin-actin complexes show that addition of G-actin to serum results in preferential formation of actin-DBP complexes, but that addition of F- actin to serum produces both gelsolin-actin complexes and DBP-actin complexes. The distinctive binding of actin monomers and polymers to these two serum proteins suggests a means by which their coordinated actions are maximized in vivo, from the standpoint of depolymerizing filaments and clearing monomers from the circulation.

scholarly journals Demonstration of alpha 1-antitrypsin by immunofluorescence on paraffin-embedded hepatic and pancreatic tissue.

1979 ◽  
Vol 27 (3) ◽  
pp. 794-796 ◽  
Author(s):  
M J McElrath ◽  
R M Galbraith ◽  
R C Allen
Keyword(s):  
Human Serum ◽  
Solid Phase ◽  
Islet Cells ◽  
Pancreatic Tissue ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Normal Human ◽  
Liver Biopsies ◽  
Alpha 1 Antitrypsin ◽  
Sepharose 4B

Studies were performed in an attempt to improve current immunohistological techniques for the demonstration of alpha 1-antitrypsin (A1AT) in formalin-fixed paraffin-embedded tissues. The unwanted fluorescence (UF) commonly occurring in such procedures was found to be effectively eliminated by immunoadsorption of A1AT antisera with human serum lacking A1AT (Pi-null phenotype) coupled in solid phase to glutaraldehyde-activated aminohexyl-Sepharose 4B. Specificity of the antisera for A1AT was established by subsequent solid phase immunoadsorption against normal human serum bound to AH-Sepharose 4B. Using these techniques, immunoreactive A1AT was demonstrated in the cytoplasm of hepatocytes in liver biopsies obtained from patients with Z and MZ serum phenotypes, and in the cytoplasm of normal pancreatic islet cells.

Cystine in Human Serum Proteins

1956 ◽  
Vol 2 (2) ◽  
pp. 65-74 ◽  
Author(s):  
Miriam Reiner ◽  
Michael X Sullivan
Keyword(s):  
Multiple Myeloma ◽  
Human Serum ◽  
Serum Protein ◽  
Serum Proteins ◽  
Normal Serum ◽  
The Other ◽  
Protein Fractions ◽  
Human Hemoglobin ◽  
Average Value ◽  
Human Serum Proteins

Abstract 1. The amount of cystine was determined in a number of serum protein fractions separated by the procedure of E. J. Cohn. In albumin, the average value was 6.40 per cent for cystine; for γ-globulin the average value was 1.94 per cent; the other fractions tested were mixtures and varied according to the different globulins present. Globin from human hemoglobin contained 1.90 per cent cystine. 2. Separated fractions from three types of multiple myeloma cases have been presented showing abnormal fractions in the α-, β-, and γregions. The percentage of cystine was determined, and except for the fraction from the "gamma" type of myelomas, which contained 6.15 per cent cystine, the anomalous protein fractions contained about the same amount found in separated fractions of normal serum.

scholarly journals Capacity of human serum to depolymerize actin filaments

Blood ◽  
1987 ◽  
Vol 70 (2) ◽  
pp. 524-530 ◽  
Author(s):  
PA Janmey ◽  
SE Lind
Keyword(s):  
Human Serum ◽  
Actin Filaments ◽  
Serum Proteins ◽  
Normal Serum ◽  
Slow Phase ◽  
Rapid Phase ◽  
Plasma Gelsolin ◽  
Preferential Formation

Abstract Human blood depolymerizes filamentous (F-)actin. The interaction of actin filaments and monomers with human serum was studied by following the kinetics and extent of the depolymerization of pyrene-labeled F- actin and by analysis of serum proteins adhering to immobilized actin monomers. In physiologic Ca2+ concentrations, the depolymerization of F- actin proceeds in two stages: a rapid phase, attributed to direct severing of filaments by plasma gelsolin, and a slow phase attributed to the binding of actin monomers to vitamin D-binding protein (DBP). Without Ca2+, only the slow phase is observed. Human serum can completely depolymerize 10 to 18 mumol/L of actin, of which approximately 5 mumol/L occurs rapidly. Depolymerization can be accounted for by the normal serum concentrations of gelsolin and DBP. Fibrin(ogen) and fibronectin, which bind actin in vitro, do not contribute to the kinetics or extent of its depolymerization. Affinity chromatography and functional assays for the presence of gelsolin-actin complexes show that addition of G-actin to serum results in preferential formation of actin-DBP complexes, but that addition of F- actin to serum produces both gelsolin-actin complexes and DBP-actin complexes. The distinctive binding of actin monomers and polymers to these two serum proteins suggests a means by which their coordinated actions are maximized in vivo, from the standpoint of depolymerizing filaments and clearing monomers from the circulation.

scholarly journals Integration of Solid-Phase Extraction and Reversed-Phase Chromatography in Single Protein-Coated Columns for Direct Injection of Bupivacaine in Human Serum

2020 ◽  
Vol 58 (6) ◽  
pp. 535-541
Author(s):  
Walaa Zarad ◽  
Heba El-Gendy ◽  
Ahmed Ali ◽  
Yasmine Aboulella ◽  
Samy Emara
Keyword(s):  
Solid Phase Extraction ◽  
Human Serum ◽  
Serum Proteins ◽  
Solid Phase ◽  
Direct Injection ◽  
Reversed Phase ◽  
Clinical Samples ◽  
Phase Extraction ◽  
Reversed Phase Chromatography ◽  
Single Protein

Abstract A rapid, reliable and precise integrated solid-phase extraction (SPE) and reversed-phase liquid chromatography method was developed and validated to determine bupivacaine in human serum using single protein-coated analytical columns. The protein-coated columns were packed with four different sorbents: TSK-ODS, LiChrosorb RP-8, LiChrosorb RP-2 and μ-Bondapak CN-bonded silica. The method involved direct injection of serum sample onto the columns for trapping of the analyte, clean-up from weakly retained serum endogenous components, as well as the final separation. The protein-coated columns operated in two different chromatographic modes. Serum proteins were extracted and cleaned up by SPE, whereas the final separation of bupivacaine was based on reversed-phase chromatography. The protein-coated TSK-ODS column resulted in more accurate peak integration and more reproducible results. A linear relationship between the concentrations of drug and peak areas was confirmed in the range of 100–2000 ng/mL. Detection and quantification limits were 24.85 and 85.36 ng/mL, respectively. The average recovery for bupivacaine ranged from 96.48% to 98.81%. The present methodology was successfully applied, with a high degree of confidence, to analyze clinical samples obtained from patient receiving 0.5% bupivacaine therapy.

INTERFERENCE BY SERUM PROTEINS WITH LH RADIOIMMUNOASSAY USING IMMUNOSORBENT

1973 ◽  
Vol 72 (2) ◽  
pp. 235-242 ◽  
Author(s):  
A. M. Reuter ◽  
J. C. Hendrick ◽  
J. Sulon ◽  
P. Franchimont
Keyword(s):  
Molecular Weight ◽  
Human Serum ◽  
High Molecular Weight ◽  
Serum Proteins ◽  
Void Volume ◽  
High Molecular Weight Proteins ◽  
Serum Fractionation ◽  
Covalently Bound

ABSTRACT The percentage of LH* bound to antibodies that have been covalently bound to cellulose is diminished in the presence of LH-free human serum and sera from various species of animals. Serum fractionation studies on Sephadex G 200 show that the greatest interference comes from the proteins eluted in the void volume i. e. the high molecular weight proteins. Specifically, the gamma M globulins and the α2-macroglobulins appear to play an important role, as demonstrated by tests in which these proteins were neutralized by gamma M and α2-macroglobulin antisera.

TESTOSTERONE BINDING BY HUMAN SERUM PROTEINS

1971 ◽  
Vol 68 (1_Supplb) ◽  
pp. S105 ◽  
Author(s):  
O. A. Lea ◽  
K. F. Støa
Keyword(s):  
Human Serum ◽  
Serum Proteins ◽  
Human Serum Proteins

STUDIES ON SULFATION FACTOR (SF) ACTIVITY OF HUMAN SERUM

1960 ◽  
Vol XXXV (III) ◽  
pp. 381-396 ◽  
Author(s):  
Sven Almqvist
Keyword(s):  
Human Serum ◽  
Normal Serum ◽  
Treatment Level ◽  
Response Curves ◽  
Normal Children ◽  
Significant Difference ◽  
Human Sera ◽  
Normal Humans ◽  
Pre Treatment ◽  
Dose Levels

ABSTRACT The sulfation factor (SF) activity of human sera has been estimated using a modification of the method of Daughaday et al. (1959). Each assay was statistically evaluated. The method had a mean precision of 0.14 and, used as an assay of GH of human serum, a sensitivity in three pituitary dwarfs of 0.1 to 0.6 μg of HGH/ml of serum. SF activity was found at all ages between 1 month and 75 years. There was a significantly lower mean SF activity below the age of half a year. Three cases of pituitary dwarfism had significantly low SF activities of sera. There was no significant difference between the SF activities of sera from untreated pituitary dwarfs and the sera from normal children below half a year of age. Dose-response curves with large volumes of sera from pituitary dwarfs and small volumes of sera from normal humans had the same slopes. Four mg of HGH prepared according to the method of Li & Papkoff (1956) resulted in a normal serum SF activity in each of the three dwarfs. A significant (P < 0.01) linear relationship was found between the concentration of SF activity of sera from these subjects and the logarithm of the dose of HGH given with dose levels of 1, 2 and 4 mg daily for three days. The decline of serum SF activity to the pre-treatment level following HGH in one dwarf suggested a half life not different from that indicated by others for growth hormone.

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