SARS-CoV-2 Neutralizing Antibody Test Cassette SARS-CoV-2 Neutralizing antibody Test Cassette NINGBO AUTRENDS INTERNATIONAL TRADE CO., LTD , https://www.metests.com
Agarose is selected and made from pure agar (agar) as a raw material. Agar is chemically a complex composed of agarose and agarose. Agar gum is a polysaccharide containing sulfate and hydroxyl groups, which has ion exchange properties, which will affect the electrophoresis and gel filtration. Agarose is a linear polysaccharide composed of alternately arranged residues of D-galactose and 3,6-anhydro-L-galactose.
Agarose forms a gel mainly by hydrogen bonding. Due to the large water content of the gel during electrophoresis (98-99%), it is approximately free electrophoresis. Because the effect of the solid support is small, the electrophoresis speed is fast and the zone is neat. Moreover, since agarose does not contain a charged group, electroosmosis has little effect, and it is a good electrophoretic material, and the separation effect is good.
Lipids in serum are present in combination with serum apolipoproteins in the form of water-soluble lipoproteins. The types and amounts of apolipoproteins contained in various lipoproteins vary, and the size of various lipoprotein particles varies greatly. Therefore, using agarose gel as a support, various lipoprotein particles can be separated in an electric field. Come.
Separation of serum proteins by agarose gel electrophoresis is simple. Serum lipoprotein is pre-stained with the lipid dye Sudan Black (or oil red, etc.). The pre-stained serum was then loaded into an agarose gel plate loading tank, and after being energized, lipoproteins were observed to move toward the positive electrode, and several zones were separated.
Normal human serum lipoprotein can appear in three zones, from the cathode to the anode, β-lipoprotein (deep), pre-beta-lipoprotein (shallow) and α-lipoprotein (slightly deeper than the pre-beta-lipoprotein). There should be no chylomicrons at the origin. Sometimes the pre-beta-lipoprotein is not shown.
Agarose forms a gel mainly by hydrogen bonding. Due to the large water content of the gel during electrophoresis (98-99%), it is approximately free electrophoresis. Because the effect of the solid support is small, the electrophoresis speed is fast and the zone is neat. Moreover, since agarose does not contain a charged group, electroosmosis has little effect, and it is a good electrophoretic material, and the separation effect is good.
Lipids in serum are present in combination with serum apolipoproteins in the form of water-soluble lipoproteins. The types and amounts of apolipoproteins contained in various lipoproteins are different, and the size of various lipoprotein particles is also very different. Therefore, using agarose gel as a support, various lipoprotein particles can be separated in an electric field. Come.
Separation of serum proteins by agarose gel electrophoresis is simple. Serum lipoprotein is pre-stained with the lipid dye Sudan Black (or oil red, etc.). The pre-stained serum was then loaded into an agarose gel plate loading tank, and after being energized, lipoproteins were observed to move toward the positive electrode, and several zones were separated.
Normal human serum lipoprotein can appear in three zones, from the cathode to the anode, β-lipoprotein (deep), pre-beta-lipoprotein (shallow) and α-lipoprotein (slightly deeper than the pre-beta-lipoprotein). There should be no chylomicrons at the origin. Sometimes the pre-beta-lipoprotein is not shown.
ã€operating】
1. 0.2 ml of the pre-stained serum serum was added with 0.2 ml of Sudan black staining solution, mixed and stained in a 37 ° C water bath for 30 minutes, and centrifuged (2000 rpm) for about 5 minutes. To remove the dye residue suspended in the serum.
2. Preparation of agarose gel plate The prepared 0.5% agarose gel was heated and melted in a boiling water bath, and the gel solution was aspirated by a pipette and poured on a glass slide, about 3 ml. Allow to stand for half an hour after solidification (expanded when hot, or put into the refrigerator for a few minutes to accelerate coagulation).
3. Draw the cut filter paper in half, and cut the sample at a distance of 2 cm from the end of the gel. Insert the capillary into the pre-stained serum. After inhaling part of the serum, take the capillary so that one end of the sample is placed against the end of the sample mouth and stop for about 3 seconds.
4, electrophoresis, put the gel plate into the electrophoresis tank, make the sample end connected to the cathode side, use four layers of filter paper or gauze to make the "leading bridge", apply to the two ends of the rubber sheet, each to hold the gel plate About 1 cm or so, the other end of the "bridge" is immersed in the buffer in the electrophoresis tank. Turn on the power, first adjust the current to 3-4mA / gel plate, electrophoresis for 10-15 minutes; then adjust the current to 6-7mA / gel plate, electrophoresis for 30-40 minutes, you can observe the separation zone.
5. If it is necessary to retain the electropherogram, the electrophoresis gel plate (with the slide) can be placed in clean water for immersion for 2 hours, and then dried in an oven (about 80 ° C).
ã€Reagents】
1. Sudan black staining solution added Sudan Black B to absolute ethanol to saturation and shake to acetylate. Filter before use.
2. The buffer was weighed for 15.4g, 2.76g and EDTA 0.29g. After adding water, add distilled water to 1000ml (pH 8.6, ionic strength 0.075) as electrode buffer.
3. The gel buffer solution was taken from 1.212 g of tris(hydroxymethyl)aminomethane, 0.29 g of EDTA, and 5.85 g of NaCl. After dissolving in distilled water, it was diluted to 1000 ml, and the pH was 8.6.
4. Agarose gel Weighed 0.45 g of agarose in 50 ml of gel buffer and added 50 ml of water. Heat to boiling in a water bath and stop heating immediately after the agarose is completely dissolved.
ã€Precautions】
1. The electrophoresis sample should be fresh fasting serum.
2. When heating and melting agar, it is necessary to prevent excessive evaporation of water. The agarose gel is used as needed to prevent the gel surface from drying and affecting the separation effect.
3. When preparing the gel plate, the agarose concentration is generally about 0.5%, the α-lipoprotein fraction is more than 1%, the β and pre-beta-lipoprotein fractions are not clear enough; the coagulation is less than 4.5%. Poor, the map is unclear.
4, the sample mouth should be the appropriate size, the edges are neat and smooth, otherwise it will affect the electropherogram.
5. If there is a shallow zone before α-lipoprotein, it can be listed as pre-alpha-lipoprotein.
[normal reference range]
--lipoprotein 20~30%
--lipoprotein 20~30%
Pre-β-lipoprotein 0~28%
Milk thistle particles (-)
Protein Technology Topics: Serum Protein Sepharose Electrophoresis
ã€principle】