Partial Purification of Urinary Erythropoietin.

Winkert, John W.; Gordon, Albert S.; Piliero, Sam J.; Medici, Paul T.

doi:10.3181/00379727-98-24041

Cited by 8 publications

(2 citation statements)

References 0 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…The cationic EIF that passed through the membrane with the permeability limited to less than a mol wt of 30,000 was retained by the membrane with the cutoff a t 10,000 and was comparable to the basic EIF fraction with a sedimentation coefficient of 1.99 S~O ,~ (9). Previous reports suggested an essential role for a basic protein associated with ESF activity (18,19), which was comparable to the cationic EGF described herein. We have demonstrated an ESF that appeared smaller than EGF (1-3), which was comparable to the anionic ESF described herein.…”

Section: Discussionsupporting

confidence: 75%

The Isolation of Erythropoiesis Regulatory Factors by an Electrofractionation Technique Combined with Selective Membrane Permeability

Lewis

Neal

Welch

et al. 1973

Experimental Biology and Medicine

View full text Add to dashboard Cite

Selective membrane permeability to a relatively small erythropoiesis stimulatory factor (ESF) was demonstrated ( 1-3). A mixture of a relatively large ESF and an ESF-generating factor (EGF) was separated by selective membrane filtration (4). This report describes the isolation of four erythropoiesis regulatory factors (ERF) , including an erythropoiesis inhibitory factor (EIF) , by combining an electrofractionation technique with selective membrane permeability. Materials and Methods.A urine concentrate of ESF, fraction I1 + I11 from a patient with paroxysmal nocturnal hemoglobinuria, was the starting material (5). The posthypoxic-polycythemic mouse was used as an assay animal (6). Results were calculated as the mean and standard error of the mean of the ESF activity of five or more mice and reported as percentage 59Fe incorporated during 48 hr and as International Units of standard B (7) ; assays with high activities were repeated at levels commensurate with linear dose-response data. Nitrogen was determined by a micro-Kjeldahl method.Fraction I1 + I11 was placed in the center section of an electrofractionation apparatus ( Fig. 1) designed and constructed for the purpose of isolating ERF. Since the buffers became progressively acidic at the anode and alkaline at the cathode during operation, the buffers chosen were 0.13 M phosphate and pH 7.9 at the anode and 5.0 at the cathode.The entire system was 0.7 M NaCl. The solutions in the anode and cathode buffer compartments were replaced every 3.5 days to 1 Supported in part by Grants FR-0061, FR-5365 and HE-12958 from the National Institutes of H 4 t h .reestablish the initial pH gradient between the compartments; during the period of 3.5 days the pH of the solution in the center section decreased from 7.4 to 7.0. Although the experiments were done at 3 t 2' and high ionic strength, enough phenol was added to the buffers to make the entire system 0.1% phenol as recommended by Lowy and Keighley (8). The chlorine and hydrogen gases evolved during electrolysis were aspirated. The solutions in the sections nearest the buffer compartments were stirred continuously. The fractions in the first anode and cathode sections were collected and replaced with fresh buffer after every 2 wk of electrophoresis through 8 wk. During this period the pH of the solution in the center section decreased to 5.9.During preliminary experiments various combinations of membranes2 were tried. Ultimately an appropriate selection of membranes was made as follows: The buffer compartments (capacity 1,500 ml each) were separated from the fraction collecting sections (capacity 250 ml each) by membranes with a molecular weight cutoff at 10,000; the first anionic fraction collecting section was separated from the second section by a membrane with a molecular weight cutoff at 20,000; the first anionic fraction collecting section was separated from the center section by a membrane with a cutoff at 30,000; the center section (capacity 500 ml) was separated from the first cationic fraction collecting sectio...

show abstract

Section: Discussionsupporting

confidence: 75%

The Isolation of Erythropoiesis Regulatory Factors by an Electrofractionation Technique Combined with Selective Membrane Permeability

Lewis

Neal

Welch

et al. 1973

Experimental Biology and Medicine

View full text Add to dashboard Cite

show abstract

“…De nombreuses publications ont trait à l'étude chez l'homme, du facteur érythropoiétique. On trouve celui-ci dans le plasma et l'urine des patients anémiques, quelle que soit l'origine de leur anémie et quel que soit leur âge; anémie pernicieuse anémie aplastique (2,73.114,168)^ thalassémie (137,176)^ anémie falciforme (73,137,176)_ ley, cémie (2,142.176) Seuls font exception les sujets anémiques atteints de néphropathies (53,98) Cette observation suggère elle aussi que le rein pourrait être le lieu de formation de l'érythropoiétine.…”

unclassified