Paper electrophoresis

1. The instrument
includes electrophoresis room and DC power supply. Common horizontal electrophoresis chamber device as shown, including two electrophoresis tank A and a sealable glass

(or corresponding material) cover B; 0.8cm)D; Rieger is a lycopic glass electrophoresis slot holder F for
filter paper
E, which can be removed from the slot, and platinum electrode D in electrophoresis slot A on both sides is connected to the external electrophoresis power supply through the groove wall by an isolated wire. The power supply is a DC power supply with a regulator, normally electrophoresis is generally 100 to 500V, and high voltage electrophoresis is generally between 500 and 10,000V.
2. Operation
(1) Electrophoretic buffer
acid buffer (pH3.0)acid (C6H8O7) H2O) 39.04gsodium oxide (C6H5Na3O7.2H2O) 4.12g, plus water 4000ml, so dissolved.(2) filter paper
take
chromatography
filter paper soaked overnight in a 1mol/L methic acid solution, the next day removed, rinsed with water to the pH of the lotion is not less than 4, put 60 degrees C
oven
drying, spare. Can be tailored according to the need to grow 27cm, width 18cm filter paper, or according to the size of the electrophoresis chamber cropped, and from the length direction of one end 5 to 8cm to draw a starting line, every 2.5 to 3cm to do a mark to prepare a sample.(3) dosing
have wet point method and dry point method. Wet point method is to cut the filter paper all immersed in theacid buffer (pH3.0), wet, take out, use the filter paper to absorb excess buffer, set the electrophoresis tank rack, so that the starting line near the yin extreme, the filter paper at both ends immersed in the buffer, and then with trace syringe precision point added to the test solution, each point 10 sl, a total of 3 points, and leave 2 blank positions. Dry point method is to test the solution point on the filter paper, blow dry, re-point, repeated several times, until the point of the specified amount of the test solution, and then spray the filter paper with a sprayer, the sample at the end of the spray wet, this method is applicable to the rare test solution.(4) electrophoresis
in the electrophoresis tank to add a moderate amount of electrophoresis buffer, immersed in platinum electrodes, plug on the electrophoresis instrument to control the power supply, adjust the voltage gradient of 18 to 20V/cm, electrophoresis about 1 hour and 45 minutes, take out, immediately blow-dry, under the ultraviolet lamp (254nm), with a pencil to draw out the purple spot position.(5) content determination
cut off the test spot and the spot location area similar to the blank filter paper, cut into fine strips, respectively,
test tube
, each precision added 0.01mol/L salt Acid solution 5 ml, shake well, place for 1 hour, with the 3rd hanging glass
fleet
filtered, can also be used natural subsidion or centrifugation method to pour the liquid, according to the provisions of each drug to determine absorption, and according to the absorption coefficient to calculate the content.