Serum Protein Electrophoresis (SPE)

Biomolecules such as proteins carry a negative or positive charge in a buffer and move toward an anode or a cathode in an electric field, which is called electrophoresis. Because of their different isoelectric points, different molecular size, shape and charge-to-mass ratio, different protein molecules have different electrophoretic mobility, and various proteins can be separated in a certain supporting medium. Commonly used electrophoresis techniques include cellulose acetate membrane electrophoresis, agarose gel electrophoresis, polyacrylamide gel electrophoresis, and immunoelectrophoresis. In addition, multiple myeloma often has M globulin bands between the beta globulin region and the gamma globulin region; double albuminemia shows a double albumin band. Basic Information Specialist classification: growth and development check classification: biochemical examination Applicable gender: whether men and women apply fasting: fasting Including items: serum β2 microglobulin (β2-MG), serum α2-macroglobulin, serum α1-microglobulin Tips: fasting 12 hours, venous blood test, specimens must be fresh blood. Before the test, you should inform the doctor about the drugs you have taken recently and the recent physiological changes. Normal value Cellulose acetate membrane electrophoresis albumin 0.61 ~ 0.71 (61% ~ 71%), α1 globulin 0.03 ~ 0.04 (3% ~ 4%), α2 globulin 0.06 ~ 0.10 (6% ~ 10%), beta globulin 0.07 to 0.11 (7% to 11%) and gamma globulin 0.09 to 0.18 (9% to 18%). Clinical significance 1, albumin reduction is found in chronic hepatitis, cirrhosis, liver cancer and so on. 2, α1 globulin (glycoprotein) increased in primary liver cancer. It is reduced in severe hepatitis, cirrhosis, and hepatic coma, and is positively correlated with albumin. The determination of α1 globulin in liver disease has reference value for judging the severity and prognosis of hepatitis. In general, the increase of α1 globulin indicates that the condition is mild, and the decrease of α1 globulin indicates that the condition is heavier. In severe liver failure, the serum content can be significantly reduced. 3. There was no significant change in the initial stage of α2 globulin viral hepatitis, and gradually increased after one week. Subacute hepatitis and acute hepatic necrosis and decompensated cirrhosis were reduced. 4, beta globulin increased in fatty liver, hyperlipidemia, nephrotic syndrome, diabetes with hypercholesterolemia, obstructive jaundice, malignant tumors. In cholestasis liver disease, its content is increased, parallel to the rise of α2 globulin. Reduced in acute and chronic hepatitis, cirrhosis, especially decompensated cirrhosis and necrotic cirrhosis decreased most significantly. 5, gamma globulin increased in chronic hepatitis, cirrhosis, liver and gallbladder disease. Typical cirrhosis can be seen in the fusion of β and γ bands to form a β-γ bridge. Changes in gamma globulin in patients with liver disease can reflect the severity of the condition. As the condition improves, its content can gradually drop to normal. If it continues to fall at a high level, it indicates that the condition is serious, the prognosis is poor, and there is a tendency to develop chronic hepatitis and cirrhosis. In addition, infection, schistosomiasis, multiple myeloma, connective tissue disease, etc. can also be elevated. Reduce the incidence of gamma globulin deficiency, partial chemotherapy patients. In addition, multiple myeloma often has M globulin bands between the beta globulin region and the gamma globulin region; double albuminemia shows a double albumin band. Low results may be diseases: high results of cholestatic jaundice may be diseases: liver cirrhosis, primary liver cancer, multiple myeloma in the elderly 1, fasting 12 hours, venous blood test, specimens must be fresh blood. 2. Before the test, you should inform the doctor about the drugs you have taken recently and the recent physiological changes. 3. Physiological elevation may occur in the following cases: (1) The increase in α1-globulin can be seen after 6 months of gestation. (2) The increase of α2-globulin can be seen in infants born 1 to 6 months, with a moderate increase in 4 to 6 months of pregnancy and a significant increase in 7 to 9 months. (3) β-globulin elevation is seen in the baby after birth, 4 to 6 months of pregnancy. (4) The increase of γ-globulin is seen after the vaccination immunization vaccine. Inspection process 1. Add the buffer to the electrophoresis tank and adjust the buffer in the tanks on both sides to make them in the same plane. 2. Preparation of cellulose acetate film: Take a cellulose acetate film (2 cm × 8 cm) and draw a horizontal line with a pencil at 1.5 cm (one side of the negative side) of the rough surface to make a dotted mark. After numbering and indicating the positive and negative electrodes, the film was immersed in the barbiturate-barbital sodium buffer solution, and after being sufficiently saturated (usually 20 min), the excess buffer was removed by sandwiching the clean filter paper. 3. The cellulose acetate film hair is attached to the electrophoresis tank holder and straightened. Pipette 3~5μl of non-hemolytic serum with a micropipette and add it along the horizontal line at the horizontal line. The sample should be kept at a certain distance from the edge of the film to avoid deformation of the protein band in the electrophoresis pattern. After the serum penetrates into the film, the film is reversed. With the light side facing up, stick it flat on the electrophoresis tank bracket, and connect the two ends of the film to the buffer with double-layer filter paper or four layers of gauze, and wait for a while. 4, turn on the power, pay attention to the positive and negative electrodes on the cellulose acetate film, do not connect wrong. Voltage 90 ~ 150v, current 0.4 ~ 0.6mA / cm (different electrophoresis required voltage, current may be different, should be flexible), summer power 45min, winter power-on time is slightly longer, about 60min, electrophoresis zone expansion about 25 ~35mm can be. 5, dyeing: After the power is completed, remove the film and directly immerse in the Lichunhong S dye solution or the amino black 10B staining solution, stain for 5 to 10 minutes (by the albumin zone), then rinse the remaining in the rinse solution. Dye until the background is colorless. 6, quantitative: 1 colorimetric method: blot the rinsed film, cut the dyed protein area into the corresponding tube, add 0.6ml/L sodium hydroxide 6ml in the albumin tube (calculate the absorbance multiplied by 2), the rest Add 3 ml to each tube, shake it several times, and place it in a 37 ° C water tank for 20 minutes to make its color leaching. Amino Black 10B staining The absorbance of each tube was read at 620 nm using a spectrophotometer, and then the contents of each tube were calculated (simultaneous blank tube control). When the Lichunhong S was dyed, the leachate was treated with 0.1 mol/L sodium hydroxide, and the amount was the same as above. After 10 minutes, add 0.6ml of 400ml/L acetic acid to the albumin tube (multiply the absorbance by 2), and add 0.3ml to the other tubes to neutralize some of the sodium hydroxide to make the color deeper. If necessary, centrifuge and take the supernatant. The absorbance of each tube was read at 520 nm by a spectrophotometer (simultaneous blank control), and then the respective contents were calculated. 2 Densitometer scanning method: A. Transparent: Aspirate the rinsing liquid on the film (to prevent the transparent liquid from being diluted to affect the transparent result), immerse the film in the transparent liquid for 2 to 3 minutes, then take it out and flatten it in a rolling manner. Clean the scratch-free glass piece (do not create bubbles), set the slide for a while, remove the excess transparent liquid, place it in an oven at a constant temperature of 90 ° C to 100 ° C, bake for 10 to 15 minutes, remove and cool to At room temperature, the protein areas transparent by this method are distinct, the film is flat, and can be directly scanned and permanently preserved (transparent with hydrogen naphthalene or liquid paraffin. The rinsed film should be dried and transparent, and the transparent film cannot be transparent) It lasts for a long time and is prone to wrinkles). B. Scanning Quantification: The transparent film is placed in a fully automatic densitometer or other densitometer dark box for scanning analysis. Not suitable for the crowd no. Adverse reactions and risks no.