Cellulose acetate electrophoresis

Hb-A2 can also be quantitated by electrophoresis. The most accurate procedures are starch block electrophoresis, cellulose acetate electrophoresis, and isoelectric focusing. 

Ammonia concentrations increased rapidly in stored blood. Blood that contained 160 103 jug/100 ml when it was collected demonstrated 665 188 jug/100 ml after 24 hours and 1181 21 /ng/100 ml in 7 days (PIO). Specimens for lipoprotein electrophoresis (cellulose acetate) can be stored at room temperature for 3 days, and in the freezer for 14 days (WIO). A second freezing and thawing made the specimen unsuitable. In... 

Most practical applications of electrophoresis in biochemistry employ some form of zonal electrophoresis, in which the aqueous ionic solution is carried in a solid support and samples are applied as spots or bands of material. Paper electrophoresis, cellulose acetate strip and cellulose nitrate strip, and gel electrophoresis are all examples of zonal... 

Chrambach this indicates that the effective protein size for gel filtration is larger than the effective size for gel electrophoresis. They concluded that this could not be accounted for by gel swelling, pH, or ionic strength effects. Biefer and Mason [36] found the constant a in Eq. (93) to be 0.93. They measured the conductance of cellulose acetate filter pads with porosities from 0.5 to 0.9 in solutions of 10 M KCl. 

Bode, H-J, The Use of Liquid Polyacrylamide in Electrophoresis III. Properties of Liquid Polyacrylamide in the Presence of Cellulose Acetate, Analytical Biochemistry 92, 99, 1979. Bowman, CN Peppas, NA, A Kinetic Gelation Method for the Simulation of Free-Radical Polymerizations, Chemical Engineering Science 47, 1411, 1992. 

Figure 50-2. Technique of cellulose acetate zone electrophoresis. A A small amount of serum or other fluid is applied to a cellulose acetate strip. B Electrophoresis of sample in electrolyte buffer is performed.

Figure 4. Cellulose acetate electrophoresis of hemoglobins. Variations in the quantities of Hb-Ag are hardly detectable (cornpare the samples 2 and 3 from top). Hb-N Baltimore is a p-chain variant in which lysyl residue in position 95 is replaced by a glutarnyl residue.

The defnon6ttLOtion 0 -chain vaAijant6 In heterozygotes Is complicated by the presence of the large amount of Hb-F. Another obstacle Is the nearly Identical electrophoretic mobilities of Hb-A and the minor Hb-Fi component. Despite these difficulties, abnormalities such as AS, SS, AC, CC, SC, and others can readily be detected using cellulose acetate electrophoresis, starch gel electrophoresis, acid agar electrophoresis, and by CM-Cellulose microchromatography to be described In a separate section. 

Hb-B0Jut 6 OK yif can best be demonstrated by either cellulose acetate or starch gel electrophoresis. The amount of Hb-Bart s can differ from 1% to over 80% dependent on the abnormality Involved. Quantitative determination Is most accurately made by CM-Cellulose or CM-Sephadex chromatography. 

The presence of Individual chains In a hemoglobin variant can also be demonstrated by electrophoresis at alkaline pH after the protein has been dissociated Into Its subunits through exposure to 6 M urea In the presence of 3-mercaptoethanol. The buffer is either a barbital buffer or a tris-EDTA-boric acid buffer, pH 8.0 - 8.6, and contains 6 M urea and 3-niercapto-ethanol. Dissociation of the hemoglobin Into subunits Is best accomplished In a mixture of 1 ml 10 g% Hb (or whole hemolysate), 4 ml 6 M urea barbital or tris-EDTA-boric acid buffer, and 1 to 1.5 ml 3-mercaptoethanol. After 30 minutes to 1 hour the sample Is subjected to cellulose acetate or starch gel electrophoresis. Each chain has a specific mobility and an alteration In electrophoretic mobility easily Identifies the abnormal chain. 

The isoenzymes can be separated by electrophoresis on cellulose acetate, and Roberts, et al ( ) have described a method whereby the separated isoenzymes are eluted and then assayed kinetically. 

CK isoenzymes have been separated by electrophoresis on cellulose acetate, agar gel, agarose and polyacrylamide gel. 

Electrophoresis on cellulose acetate strips (Sepraphore III, Gelman Instrument, Ann Arbor, MI) was done in the conventional manner [12] in order to obtain a comparative electrophoretic mobility of non-adsorbed albumin. For this purpose, BSA-BSA (2.5 w/v) was deposited on the cellulose acetate paper twice in volumes of 10 ul each. Electrophoresis was attain performed in the Gelman Chamber with Pt electrodes at 20°C (see Table 3) After completion, the strips were stained with Ponceau S protein stain (Gelman Instruments) and washed with 5 acetic acid. The stained cellulose acid strips were subsequently cut into 3 mm wide pieces which were monitored for protein content y-count-ing. 

Influence of the Surface Concentration of BSA. Compared to the corrected moving boundary electrophoretic mobility of BSA in solution, the mobility of BSA adsorbed onto glass is considerably faster at all ionic strengths at 1.96 pg/cm2 and somewhat faster at lower ionic strengths 1.38 pg/cm2. However, at lower adsorption densities (1.05 and 0.64 pg/cm2), the adsorbed BSA moves more slowly in the applied electric field than BSA in moving boundary electrophoresis under otherwise identical conditions, and at the lowest surface adsorption (0.64 pg/cm2) the mobility of the adsorbed BSA are even somewhat slower than in cellulose acetate gel at all conditions of ionic strength investigated. 

TABLE 4 Comparison of the adsorbed BSA electrophoresis results of Tables 1 and 2, with cellulose acetate electrophoresis (Table 3) and moving boundary electrophoresis [6], extrapolated to 30°C (as in Tables 1 and 2) and to the appropriate ionic strengths (Tables 1-3)... 

Traditional electrophoresis paper, cellulose acetate or polymeric gels used as a supporting medium for the electrolyte solution enclosed tank with electrodes and buffer reservoirs dc power supply. 

Electrophoresis has also been employed to separate neomycin from analytically-interfering substances such as proteins. Hence Brammer and Hemsonl82 have determined the neomycin content of blood serum. Neomycin was separated from the serum proteins by electrophoresis on cellulose acetate and assayed colorimetrically following elution from the support. 

Procedure 11.7 Location of proteins after electrophoresis on cellulose acetate... 

Electrophoresis agarose, poly- Protein composition acrylamide gel, cellulose acetate... 

Concentration of the urine of patients with suspected paraproteinemia for electrophoresis need not involve the purchase of any extra equipment, and highly satisfactory electrophoretograms on cellulose acetate mem-... 

The immunoelectrophoretic results should be interpreted in conjunction with interpretation of cellulose acetate electrophoresis of both the serum and urine from patients who are suspected of having multiple myelomatosis. Indeed, it should be mentioned that no single pattern of immunoelectrophoretic precipitin line is typical of patients with multiple myelomatosis, and it can be said that these vary from patient to patient and must be interpreted with care. Frequently it may be essential to repeat... 

Polyacrylamide gel electrophoresis (PAGE) and cellulose acetate membrane electrophoresis (CAME) were applied to distinguish escolar and oilfish from 27 commercial fish based on muscle protein differences (Ochiai et al., 1984). Myogen fractions from the muscles were subjected... 

W5. Windisch, R. M., and Bracken, M. M., Cerebrospinal fluid proteins Concentration by membrane ultrafiltration and fractionation by electrophoresis on cellulose acetate. Clin. Cherru 16,416-419 (1970). 

Other supports in electrophoresis are agarose gels, paper, or celluloses acetate. 

Before discussing the various derivatives that have been used, it should be remembered that oligosaccharides are often obtained by chromatography on paper or on cellulose columns, and this may cause them to become contaminated by xylan this may be eliminated by extraction of the crude fractions with hot ethanol.474 For similar reasons, glucose that is really extraneous may be detected in fractions separated by electrophoresis on cellulose acetate.475 Gas-liquid chromatography was used to show the presence in pituitary glyco-... 

Recently, microchip electrophoresis was applied to GAG analysis using ethidium bromide as a fluorescent dye. In particular, separation times were reduced to 150 s, while sensitivity remained comparable to that of conventional electrophoretic methods that rely on cellulose acetate membranes [47]. 

Bell, J. W. and Stone, W. K. 1979. Rapid separation of whey proteins by cellulose acetate electrophoresis. J. Dairy Sci. 62, 502-504. 

West, D. W. and Towers, G. E. 1976. Cellulose acetate electrophoresis of casein proteins. Anal. Biochem. 75, 58-66. 

Electrophoresis on cellulose acetate strips has also been used for the rapid resolution of whey proteins (Bell and Stone 1979). Samples of a 10 1 concentrate of whey are applied to cellulose acetate strips which have been saturated with Tris-barbiturate buffer, pH 8.6, ionic strength 0.097, and the electrophoresis is performed at 225 V for 1 hr. This procedure separates not only the major whey proteins but also their genetic variants. 

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