Method for marking antibody by using alkaline phosphatase
A technology for labeling antibodies and phosphatases, applied in the field of biomedicine, can solve problems such as complex operations, achieve the effects of optimizing oxidation time, simplifying operations, and improving labeling effects
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Embodiment 1
[0025] A) Dissolve 5 mg of alkaline phosphatase in 0.5 ml of NaAc solution with a concentration of 0.05 M and a pH value of 5.6.
[0026] B) Add 0.5ml of 0.06M sodium periodate solution to the enzyme solution in step A, and oxidize at a room temperature of 4 degrees Celsius for 20 minutes.
[0027] C) Add 0.5 ml of ethylene glycol-NaCl solution to the oxidized enzyme solution. The formula of ethylene glycol-NaCl solution is: Weigh 22g NaCl, add 2.3ml ethylene glycol, and add water to 100ml to prepare the solution, which can be stored at room temperature for a long time.
[0028] D) Add 1.5 ml of ice-cold absolute ethanol to the solution in step C, mix well, let stand at room temperature at -20° C. for 1 hour, and centrifuge at 4000 g for 10 min to obtain the oxidized enzyme.
[0029] E) Dissolve the enzyme obtained in the precipitation step D with 0.5 ml of carbonate buffer solution with a concentration of 0.5 M and a pH value of 9.6, add 0.1 ml of glutaraldehyde, and mix wel...
Embodiment 2
[0037] A) Dissolve 5 mg of alkaline phosphatase in 0.5 ml of NaAc solution with a concentration of 0.05 M and a pH value of 3.6.
[0038] B) Add 0.5ml of 0.06M sodium periodate solution to the enzyme solution in step A, and oxidize at a room temperature of 4 degrees Celsius for 20 minutes.
[0039] C) Add 0.5 ml of ethylene glycol-NaCl solution to the oxidized enzyme solution. The formula of ethylene glycol-NaCl solution is: Weigh 22g NaCl, add 2.3ml ethylene glycol, and add water to 100ml to prepare the solution, which can be stored at room temperature for a long time.
[0040] D) Add 1.5 ml of ice-cold absolute ethanol to the solution in step C, mix well, let stand at room temperature at -20° C. for 1 hour, and centrifuge at 4000 g for 10 min to obtain the oxidized enzyme.
[0041] E) Dissolve the enzyme obtained in the precipitation step D with 0.5 ml of carbonate buffer solution with a concentration of 0.5 M and a pH value of 9.6, add 0.1 ml of glutaraldehyde, and mix wel...
Embodiment 3
[0049] A) Dissolve 5 mg of alkaline phosphatase in 0.5 ml of NaAc solution with a concentration of 0.05 M and a pH value of 5.6.
[0050] B) Add 0.5ml of 0.1M sodium periodate solution to the enzyme solution in step A, and oxidize at a room temperature of 4 degrees Celsius for 20 minutes.
[0051] C) Add 0.5 ml of ethylene glycol-NaCl solution to the oxidized enzyme solution. The formula of ethylene glycol-NaCl solution is: Weigh 22g NaCl, add 2.3ml ethylene glycol, and add water to 100ml to prepare the solution, which can be stored at room temperature for a long time.
[0052] D) Add 1.5 ml of ice-cold absolute ethanol to the solution in step C, mix well, let stand at room temperature at -20° C. for 1 hour, and centrifuge at 4000 g for 10 min to obtain the oxidized enzyme.
[0053] E) Dissolve the enzyme obtained in the precipitation step D with 0.5 ml of carbonate buffer solution with a concentration of 0.5 M and a pH value of 9.6, add 0.1 ml of glutaraldehyde, and mix well...
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