Cellulose Acetate Electrophoresis of Hemoglobin.

The electrophoresis on cellulose acetate membrane is most widely used because of its simplicity, and is without the use of any sophisticated instrument other than electrophoresis apparatus and the cellulose acetate strip. Here we describe a modified version of cellulose acetate membrane electrophoresis for hemoglobin separation from blood sample. Sharp, clear bands without tailing effects can be obtained with this method. The method and apparatus described here would be appropriate to separate protein fractions under 1 h at voltages up to 60 V/cm measured between the electrodes.

Preliminary Evaluation of a Point-of-Care Testing Device (SickleSCAN™) in Screening for Sickle Cell Disease.

Sickle cell disease affects about 150,000 births annually in Nigeria. Early diagnosis is hampered by factors such as centralized and urban localization of laboratories, high cost of diagnostic equipment and inadequate skilled manpower to operate them. The need for a low-cost, portable, easy-to-use diagnostic test for sickle cell disease is critical, especially in resource-poor countries. In this study, we evaluated the performance characteristics of a novel point-of-care testing device (SickleSCAN™), and its acceptability and feasibility, as a possible screening tool for sickle cell disease. In the first phase, we assessed the performance characteristics of SickleSCAN™ by evaluating 57 subjects comprising both children and adults attending a primary health center, for Hb SS (ßS/ßS; HBB: c.20A>T), Hb SC (ßS/ßC; HBB: c.19G>A) and Hb AS (ßA/ßS) using SickleSCAN™, cellulose acetate electrophoresis (CAE) and high performance liquid chromatography (HPLC). Performance characteristics such as diagnostic sensitivity and specificity were compared to HPLC as a standard method. We subsequently undertook a second phase wherein the acceptability and feasibility of the device for sickle cell disease screening, was evaluated using semi-structured and structured questionnaires among 197 healthcare personnel and 221 subjects, respectively. Sickle cell disease was carried by 3.4% of the subjects. The diagnostic sensitivity, specificity and test efficiency of SickleSCAN™ for sickle cell disease (Hb SS and Hb SC), were 100.0, 98.2 and 98.2%, respectively. Findings from this study showed SickleSCAN™ to be a viable screening tool that can easily be applied in community-based screening for early diagnosis of sickle cell disease with little expertise and low cost.

A simple cellulose acetate membrane-based small lanes technique for protein electrophoresis.

Combining electrophoresis with a cellulose acetate membrane-based technique, we developed a simple and low-cost method, named cellulose acetate membrane-based small lanes (CASL), for protein electrophoresis. A home-made capillary plotter controlled by a 3D moving stage was used to create milli-to-micro channels by printing poly(dimethylsiloxane) on to a hydrophilic cellulose acetate membrane. In the hydrophilic channels, 5 nL protein mixture was separated on the basis of electro-migration under an electric field. Compared with polyacrylamide gel electrophoresis (PAGE), CASL resulted in higher protein signal intensity for separation of mixtures containing the same mass of protein. The platform was easily fabricated at low cost (approx. $0.005 for each 1-mm-wide channel), and separation of three protein mixtures was completed in 15 min. Both electrophoresis time and potential affected the separation. Rather than chromatographic separation, this method accomplished application of microchannel techniques for cellulose acetate membrane-based protein electrophoresis. It has potential in proteomic analysis, especially for rapid, low-cost, and low-volume sample analysis in clinical diagnosis.

[The comparative analysis of electrophoretic fractionating of blood serum proteins in diagnostics of multiple plasma cell myeloma].

The choice of technology of electrophoretic fractionating of blood serum proteins is determined, besides the analytical characteristics, by its economic component. The electrophoresis technologies developed by the R&D production facility "Astra" (Russia) and the firm "PZ Cormay S.A." (Poland) are compared from a viewpoint of applicability in routine laboratory, practice and diagnostics of multiple plasma cell myeloma in particular. It is established that under the comparable economic, "consumer" and analytic characteristics of technologies in the diagnostic process the application of the technology in agarose gel ("PZ Cormay S.A.") is more preferable.

Reuse of cellulose acetate membrane strips for protein and haemoglobin electrophoretic analysis.

The exorbidant cost of electrophoretic analysis, many a times becomes the major limitation factor. As most of the clinical laboratories import the cellulose acetate membrane which costs 5 to 6 U.S. $ that is Rs. 180/- to Rs. 210/-, the final cost goes upto Rs. 350/- to Rs. 500/-. We have observed that CAN strips can be effectively reused 6 to 7 times for subsequent electrophoretic analysis.

Caracterización de formas macroenzimáticas de la fosfatasa alcalina: descripción de un nuevo método / A new method for the characterization of alkaline phosphatase macroenzymes

Se describe un nuevo método para la caracterización de macroenzimas de la fosfatasa alcalina. El método combina una separación previa de las formas isoenzimáticas por gel filtración en capa fina de Sephadex G-200, y el posterior revelado de la correspondiente actividad enzimática in situ. Entre otras ventajas, este sistema no sólo separa adecuadamente las formas macroenzimáticas, sino que mantiene todas las isoenzimas en estado nativo, a diferencia de las técnicas con desnaturalizantes, por lo que es posible su detección y eventual aislamiento. Permite además la resolución simultánea de un gran número de muestras, así como la inclusión de controles de distinto peso molecular (PM) dentro de una misma corrida, lo que facilita en forma significativa la posterior interpretación del perfil obtenido, aportando una gran ventaja con respecto a la técnica de gel filtración en columna. El método propuesto se presenta como una técnica relativamente simple y rápida para el screening de macroenzimas en el laboratorio clínico

Application of the monodimensional electrophoresis for the separation of glycosaminoglycans on Cellogel cellulose acetate strip.

A monodimensional electrophoretic method for the separation of glycosaminoglycans on Titan III Zip Zone cellulose acetate plate based on their different electrophoretic mobilities in barium acetate and different solubilities in ethanol was applied to the Chemetron electrophoretic equipment. Improved timing of individual steps of electrophoretic run, additional cooling and pressure must be introduced for optimal separation of glycosaminoglycans mixture (dermatan sulphate, heparan sulphate, hyaluronic acid, chondroitin-4-sulphate, chondroitin-6-sulphate and keratan sulphate) resulting in five well separated sharp bands. By all these changes in the original procedure of Hopwood and Harrison, the separation of chondroitin-4-sulphate and chondroitin-6-sulphate was not achieved. The modified procedure on Cellogel strip is suitable for the screening of mucopolysaccharidoses.

[Apparatus for electrophoresis on cellulose acetate films]. / Pribor dlia élektroforeza na plenkakh iz atsetata tselliulozy.

Type EFPA-30 apparatus for electrophoresis on cellulose acetate films was developed at the Specialized Design Office for Biophysical Equipment of the Moscow Research and Production Association BIOFIZPRIBOR. Three autonomic separation cameras are united in one block, that permits separation of 12, 24, and 36 samples; the apparatus is supplied with time transducer, acoustic and visual indication. Clinical trials of the apparatus were carried out at the Institute of Biological and Medical Chemistry of the USSR Academy of Medical Sciences, its commercial production is to be started. The apparatus is intended for protein separation and may be used for studies of hemoglobins, serum proteins, lipoproteins, enzymes, and other biologic compounds. Methods for hemoglobin and serum protein separation are presented.

An improved applicator system for cellulose acetate electrophoresis.

An improved sample application system for cellulose acetate electrophoresis is described. This system features 30 interchangeable wells allowing sample reordering for side-by-side comparison of closely spaced electromorphs.

[Electrophoretic separation of lipoproteins on Vladipor membranes]. / Elektroforeticheskoe razdelenie lipoproteidov na membranakh "Vladipor".

Vladipor cellulose acetate membranes of various make for electrophoretic separation of lipoproteins are compared, the procedure of the membrane treatment and lipoprotein fractionation is described. Electrophoretic fractionation of human blood serum lipoproteins has been found most effective with the use of the MPA-MA Nos. 4 and 5 membranes.

Counterflow immunoisotachophoresis on cellulose acetate membranes.

Discontinuous electrophoresis on cellulose acetate membranes with the use of 0.06 M Tris-HCl (pH 6.7) as the leading electrolyte and 0.012 M Tris-beta-alanine (pH 8.6) as the terminating one results in concentration of the proteins present in the system on the Cl-/beta-alanine- boundary. If the antigen solution is placed in a "pocket" ahead of the moving boundary, a counterflow to the cathode arises due to electroendosmosis. At constant voltage the migration rate of the boundary drops and that of electroendosmosis does not change until they become equal. In such a stationary position, the antigen-containing solution is passing through the Cl-/beta-alanine- boundary to the cathode, while all the proteins are completely "absorbed" on the boundary as highly concentrated bands. Addition of ampholytes to the antigen solution contributes to the isotachophoretic separation of a protein mixture on the strip. The concentrated and separated antigens can be revealed by immunofixation, immunodiffusion, or crossed immunoelectrophoresis in gel. The technique is approximately 100 times more sensitive compared to the usual immunodiffusion and immunoelectrophoresis methods on cellulose acetate membranes, and is applicable to the detection of trace amounts of antigens in the urine, liquor, amniotic fluid, tears, and other biological fluids with low protein contents.

[Computer-assisted findings on protein electrophoresis on cellulose acetate film]. / Rechnerunterstützte Befundung von Eiweisselektrophoresen auf Celluloseacetatfolie.

Until a short time ago efficient computer assisted reporting of electrophoretograms was not possible owing to the lack of appropriate technology. By integrating a fully mechanized electrophoresis system into the laboratory data processing system of a centralized institute of clinical chemistry, the separation and the interpretation of results can be performed for 300 samples per day. The interpretation is based upon available patient data (specified clinical diagnoses, previous results), calculated fractions and analysis of the electrophoretic curve in the area of albumin and the beta- and gamma-globulin fraction. In this way it is possible to classify hereditary bisalbuminaemias and to detect transient bisalbuminaemias and monoclonal gammopathies. Diagnostic indications of specific dysproteinaemias and defect proteinaemias are printed on the report form if individual serum protein fractions and the total protein content are changed specifically. The results of the different electrophoretic fractions, the total protein and a set of possible alterations of the curve are numerically coded according to the scaled biochemical ranges in five definite sections or to a decision matrix (alteration 'given' or 'not given'), respectively. The composed 'result patterns' are matched with preassigned 'reference patterns' stored in the computer file. The procedure is efficient and adaptable to other areas of analysis.

A note of caution on the use of the LD ratio in myocardial infarction.

Different LD1/LD2 ratios for both normal and pathological samples were obtained with two different reagent kits. These ratio differences are due to the substrate formulation and not to the electrophoresis separation technique. In 45 patients with abnormal serum "cardiac" enzymes, results obtained by the two kit methods showed 17% and 50% "flipped ratio" abnormalities as against 78% and 67% abnormalities when the ratios were compared against appropriate reference ranges. The improper use of the LD ratio test is probably the main cause for its diminished diagnostic usefulness.

Two-dimensional spectroscopy of electrophoretic gels.

Optical techniques are described which permit one to analyze two-dimensional electrophoretic gels in a fashion which is analogous to the one-dimensional spectroscopy of solutions. In the methods described, an electrophoretic gel is irradiated with monochromatic light and isozyme patterns are detected by the absorption of light or the fluorescent emission of light. The system described can both generate and detect monochromatic light in a range from 200 to 1100 nm. Without the use of histochemical stains, several isozymes have been visualized by purely optical means. Five methods for the visualization of lactate dehydrogenase and five methods for the demonstration of trypsin isozymes are described. In addition, general methods have been formulated for hydrolases and oxidases. Gel spectroscopy should permit the investigation of a wide range of new isozymes.