Preparation method of microporous reaction plates, kit and detection method of kit

A technology of microwell reaction plate and kit, which is applied in the field of multiple detection of thyroid function, can solve the problems of insufficient reconstitution, confusion of reagent types, cumbersome operation, etc., achieve easy detection automation, realize detection automation, and simple experimental process Effect

Pending Publication Date: 2019-02-05
广州俊通生物科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] (1) At present, most of the standard products and tracers need to be reconstituted 30 minutes in advance before the test, and there are 7 bottles of standard products and tracers for each test item, and 35 bottles of reagents need to be reconstituted for 5 items, which not only wastes time, it is also easy to cause cross-contamination, and abnormal experimental results caused by insufficient reconstitution and inconsistency;
[0007] (2) The combined detection of multiple items of existing microplate reagents needs to be combined, which is not only cumbersome to operate, but also easy to cause confusion due to the large number of reagents;
[0008] (3) Because the tracer working solution needs to be introduced into the container when preparing, it is easy to cause pollution;
[0009] (4) Due to the need for surplus in the liquid addition process, the actual preparation of the tracer working solution is more than the theoretical dosage, and the surplus of the automation equipment is more, so the tracer loading provided by the manufacturer is often more than the actual dosage. The equipment is higher, and the excess liquid in each experiment is discarded, resulting in waste of reagents

Method used

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  • Preparation method of microporous reaction plates, kit and detection method of kit
  • Preparation method of microporous reaction plates, kit and detection method of kit
  • Preparation method of microporous reaction plates, kit and detection method of kit

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Effect test

Embodiment 1

[0057] A method for preparing a microporous reaction plate, wherein the microporous reaction plate is provided with tracer microspheres inside, and the microporous reaction plate is used for a multiple detection kit of thyroid function, and the microporous reaction plate is made by the following steps :

[0058] Step 1), preparing the microporous reaction plate of the porous microplate blank plate into a coated plate;

[0059] Step 2), preparing tracer microspheres, adding 1 mg to 3 mg of the target antigen (antibody) into 1 mg of Eu3+-N2-[P-isocyanic acid-benzyl]-diethylenetriaminetetraacetic sodium and mixing, 25 Reaction at ℃ for 24 hours; 3 mg of target antigen or antibody was used in this example;

[0060] Step 3), the reaction solution was eluted with a Sephadex G-50 column (1×40cm) with 50mmol / L, pH 7.8 Tris-HCl buffer to separate tracer conjugates and free Eu3+;

[0061] Step 4), using a fully automatic partial collector 1ml / tube for liquid collection;

[0062] Step...

Embodiment 2

[0075] A test kit, which includes reconstitution reagents, concentrated washing solution, enhancement solution, standard products, quality control products and the above-mentioned microporous reaction plate.

[0076] The reconstitution reagent is to add 1.0g / L~20.0g / L bovine serum albumin to a 50mmol / L, pH7.8 Tris-HCl solution containing 0.01g / L disodium edetate . In the present embodiment, add the bovine serum albumin of 10g / L

[0077] Wherein, the concentrated lotion is 1.0ml / L-10.0ml / L Tween-20 added to Tris-HCl buffer solution containing 0.385mol / L NaCl, 0.124mol / L, pH7.8. Add 9ml / L Tween-20 in this example.

[0078] Wherein, the enhancement solution is to add 1-5 mg / L β-naphthoyl trifluoroacetone, 15-30 mg / L's tri-n-octyl phosphine oxide. In this example, 4 mg / L β-naphthoyl trifluoroacetone and 20 mg / L tri-n-octylphosphine oxide were added.

Embodiment 3

[0080] The present invention also provides a detection method of the kit, comprising the following steps: step 1) equilibrating the microwell reaction plate strips required by the reagent to room temperature;

[0081] Step 2) Mix the above-mentioned concentrated washing solution and deionized water at a ratio of 1:20 to form a working washing solution;

[0082] Step 3) Reconstitute the standard and quality control products with purified water according to the marked volume;

[0083] Step 4) Add 100-200 μl of reconstitution reagent to each well;

[0084] Step 5) Then add 25-100 μl of standard substance, quality control substance or specimen to be tested in sequence; shake and incubate the reaction system at 37°C for 10-30 minutes;

[0085] Step 6) Wash the plate 4-6 times with the above-mentioned working washing liquid, and pat dry;

[0086] Step 7) Add 50-200 μl enhancement solution to each well, shake slowly for 5 minutes, and perform fluorescence counting in the fluorescen...

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Abstract

The invention relate to a preparation method of microporous reaction plates, a kit and a detection method of the kit. The detection method of the kit comprises the follow steps that (1) microporous reaction plates required by a reagent are balanced to room temperature; (2) concentrated washing liquid and deionized water are mixed according to a ratio of 1: 20 to form working washing liquid; (3) astandard substance and a quality control substance are redissolved with purified water according to the marked volume; (4) 100-200 [mu]l of a redissolution reagent are separately added into each hole;(5) then 25-100[mu]l of the standard substance, the quality control substance or specimen to be tested are sequentially added; oscillating incubation is carried out for 10-30 minutes; (6) the platesare washed for 4-6 times with the working washing liquid, pat dry is carried out; and (7) 50-200 [mu]l of enhancement liquid is added to each hole, slow oscillating is carried out for 5min, and fluorescence counting is carried out in a fluorescence reading device. The method reduces the risk of misoperation and pollution caused by the introduction of a preparation container; and after a tracer ismanufactured into microspheres, the microspheres are placed in the microporous reaction plates, so that the loading amount does not need to be surplus, the waste of the reagent is reduced, and the cost of the reagent is reduced.

Description

technical field [0001] The invention relates to the technical field of multiple detection of thyroid function by time-resolved immunofluorescence assay technology, in particular to a preparation method of a microporous reaction plate, a kit and a detection method of the kit. Background technique [0002] Time Resolved Immunofluorometric Assay (TRIFMA) is a new type of ultra-micro immunoassay technology that came out in the early 1980s. Fluorescence time-resolved technology is a fluorescence spectroscopy technique that uses the difference in fluorescence lifetime of fluorescent emitters to separate their fluorescence. Use lanthanides as tracers instead of radioactive isotopes, and use lanthanides with bifunctional groups to label antigens or antibodies. When the dissociated lanthanides form a new chelate with a special amplification solution, Its fluorescence is significantly enhanced and persistent. The method of delayed measurement can eliminate the background fluorescenc...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/64
CPCG01N21/6408G01N21/6428
Inventor 李婉君
Owner 广州俊通生物科技有限公司
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