Serum lipoprotein and serum lipoprotein electrophoresis
Lipoprotein electrophoresis is mainly used for the classification of hyperlipoproteinemia, and it is also helpful to understand the blood lipid status of coronary heart disease and better guide clinical diagnosis and treatment.Basic Information
Specialist classification: cardiovascular examination classification: biochemical examination
Applicable gender: whether men and women apply fasting: fastingAnalysis results:
Found in low-lipoproteinemia and so on.
Very low density lipoprotein: 0.06-0.3g/L
Low density lipoprotein: 0-7g/L
High-density lipoprotein (male): 0.41-0.63g/L
High density Zheng?.42-0.68g/L
Found in primary hyperlipidemia and so on.
Cellulose acetate membrane electrophoresis
Milk thistle particles 0g / L (0mg / dl);
Very low density lipoprotein 0.06 ~ 0.3g / L (6 ~ 30mg / dl);
Low density lipoprotein <7g / L (<700mg / dl);
High density lipoprotein:
Male 0.52 ± 0.11g / L (43 ± 18mg / dl);
Female 0.55 ± 0.13g / L (47 ± 2mg / dl).
1, increased: seen in primary hyperlipidemia.
2, lower: seen in low-lipoproteinemia.High results may be diseases: hyperlipidemia, hyperlipoproteinemia type II, coronary heart disease considerations
1. Sample: Lipoprotein can be isolated from fresh, unfrozen serum. Plasma is not suitable for lipoprotein electrophoresis because a fibrinogen band will appear.
2. Lipoprotein profiles with clear separation components are prerequisites for accurate interpretation. If these conditions are well met, the lipoprotein electrophoresis and the reference method (ultracentrifugation) can be quite consistent. The precision of each component is different, and it is estimated by the coefficient of variation, which is generally less than 5%.
3. Occasionally, alpha-lipoproteins will disappear and/or a narrow, alpha-lipoprotein band will appear. This is because free fatty acids are in. Accumulation on alpha-lipoproteins causes these particles to have equal charge. As the density of the α-zone increases, the result is high. Compared to precipitation techniques, quantitative lipoprotein electrophoresis is costly.Inspection process
Immediately after venous blood collection, the test operation:
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.
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 and scratch-free glass slides (do not create bubbles), erect the slides for a while, remove excess transparent liquid, place in an oven at a constant temperature of 90-100 ° C, bake for 10-15 min, remove and cool to 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 stored. Long, and easy to wrinkle). 2 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
Inappropriate people: Generally there are no people who are not suitable.Adverse reactions and risks
1, local subcutaneous hemorrhage: after the blood collection should be pressed for a sufficient time, especially those with bleeding tendency, so as not to cause subcutaneous oozing and bruising due to no blood coagulation.
2, infection: attention to aseptic operation during venous blood collection, so as not to cause local infection.