Ratio immunoassay method with commercial magnetic beads as internal standard
An immune analysis and ratio technology, applied in the field of immune analysis, can solve the problems that hinder the wide application of SI, and achieve the effect of simple method, avoiding inaccuracy and improving accuracy
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Embodiment 1
[0050] S1: Different amounts of CEA antibodies were incubated with 1 mL of AuNPs solution for 40 minutes.
[0051] S2: Measure the absorbance of the above mixed solution.
[0052] S3: the Ab 2 - After the AuNPs were purified by centrifugation, 300 μL of 3 M NaCl solution was added and mixed with it.
[0053] S4: Observe the color and state of the above solution.
[0054] Experimental results:
[0055] This experiment is to optimize the optimal dosage of the antibody modified gold nanoparticles of the present invention.
[0056] Wherein, the reaction conditions of this experiment are as described in Example 1.
[0057] Antibody dosages were designed to be 0, 1, 3, 5, 7, 10, 13, 15, 17, and 20 μg, respectively.
[0058] When the amount of added antibody is greater than 10 μg, Ab 2 - The absorbance of the AuNPs solution remains basically unchanged. This indicates that when the amount of antibody is about 10 μg, it is enough to complete the modification of AuNPs. And when ...
Embodiment 2
[0060] S1: Preparation of Ab 1 -MPs.
[0061] S2: Gold nanoparticles were incubated with 13 μg antibody for 40 minutes at room temperature to prepare Ab 2 -AuNPs, and blocked with BSA.
[0062] S3: Take 40 μL of diluted Ab 1 -MMP and 100 μL of fixed concentration of CEA were mixed evenly, and incubated with shaking at 37 ºC and 1500 rpm for a certain period of time.
[0063] S4: Wash the magnetic bead complex twice with 50 μL PBST solution. Then, add 50 μL of diluted Ab to the sample 2 -AuNPs, shake and incubate at 37 ºC, 1500 rpm for 1 h.
[0064] S5: Finally, after washing, the magnetic bead complex was reacted with 50 μL aqua regia for 20 min, and the solution was diluted to 8 mL with ultrapure water. Finally, the solution was collected by high-resolution inductively coupled plasma mass spectrometry 197 Au and 57 Fe signal and calculate the ratio of the two.
[0065] Experimental results:
[0066] This experiment is the Ab in the preferred invention 1 - Optimal r...
Embodiment 3
[0071] S1: Preparation of Ab 1 -MPs.
[0072] S2: Gold nanoparticles were incubated with 13 μg antibody for 40 minutes at room temperature to prepare Ab 2 -AuNPs, and blocked with BSA.
[0073] S3: Take 40 μL of diluted Ab 1 -MMP and 100 μL of fixed concentration of CEA were mixed evenly, and incubated at 37 ºC and 1500 rpm for 1 h with shaking.
[0074] S4: Wash the magnetic bead complex twice with 50 μL PBST solution. Then, add 50 μL of diluted Ab to the sample 2 -AuNPs, shake and incubate for a certain period of time under the conditions of 37 ºC and 1500 rpm.
[0075] S5: Finally, after washing, the magnetic bead complex was reacted with 50 μL aqua regia for 20 min, and the solution was diluted to 8 mL with ultrapure water. Finally, the solution was collected by high-resolution inductively coupled plasma mass spectrometry 197 Au and 57 Fe signal and calculate the ratio of the two.
[0076] Experimental results:
[0077] This experiment is the Ab in the preferred ...
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