Simultaneous purification of human albumin and hemoglobin for use as environmental biomarkers.
A combination of known biochemical techniques (binding of bromocresol green to human albumin, nondenaturing electrophoresis and electroelution) have been utilized in a novel purification of human albumin. This paper reports (a) purification of intact, nondenatured human albumin, (b) simultaneous purification of albumin and hemoglobin from a human plasma and red blood cell lysate mixture, (c) development of a purification method for the two most commonly employed protein biomarkers in human environmental epidemiology and (d) demonstration of the general technique of protein purification of colored ligand-protein complexes by nondenaturing electrophoresis. The noncovalent binding between a protein (albumin) and a colored ligand (bromocresol green) is the biochemical characteristic exploited in this novel purification scheme. This general purification method may be useful for other colored ligand or fluorescent ligand binding proteins. For small-scale electrophoresis, the amount of protein isolated, percentage yield and protein purity (estimated by SDS-PAGE) were 270 micrograms, 80% yield and > 99% purity for albumin and 217 micrograms, 54% yield and > 99% purity for hemoglobin, respectively. For large-scale electrophoresis the comparable data was 38.3 mg, 57% yield and 98% purity for albumin and 17.2 mg, 57% yield and 99% purity for hemoglobin, respectively.
Kitchin
K T
KT
Environmental Carcinogenesis Division, U.S. Environmental Protection Agency, Research Triangle Park, NC, USA.
Brown
J L
JL
D015415:Biomarkers / Q000032:analysis; D001961:Bromcresol Green / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004784:Environmental Monitoring; D004912:Erythrocytes / Q000737:chemistry*; D006454:Hemoglobins / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification*; D006801:Humans; D010949:Plasma / Q000737:chemistry*; D012709:Serum Albumin / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification*
D015415:Biomarkers; D006454:Hemoglobins; D012709:Serum Albumin; D001961:Bromcresol Green