CN111855997A - Novel immunohistochemical three-in-one reagent formula - Google Patents

Abstract

The invention relates to a novel immunohistochemical three-in-one reagent formula, belonging to the field of immunohistochemistry; the composite material consists of the following raw materials in mass number: 0.1-0.4g Triton X-100; 1ml-4ml hydrogen peroxide; 0.5ml-2ml goat serum; 0.1g-0.4g BSA; 0.01g to 0.04g of tween-20; 0.001g-0.01g EDTA; 0.1g-0.4g PC300 was dissolved in 17ml PBS, pH 7.4. The invention effectively shortens the immunohistochemical time and does not influence the immunohistochemical experiment effect.

Description

Novel immunohistochemical three-in-one reagent formula

Technical Field

The invention belongs to the field of immunohistochemistry, and particularly relates to a novel immunohistochemical three-in-one reagent formula.

Background

Immunohistochemistry is a method of determining the antigens (polypeptides and proteins) in tissue cells by applying the basic immunological principle, namely the principle of antigen-antibody reaction, namely the specific combination of antigens and antibodies, and developing color development agents (fluorescein, enzyme, metal ions and isotopes) of labeled antibodies through chemical reaction, and performing positioning, qualitative and relatively quantitative researches on the antigens, which is called immunohistochemistry (immunohistochemistry) or immunocytochemistry (immunocytochemistry).

Immunohistochemistry makes use of this principle. Firstly, a certain chemical substance in the tissue or cell is extracted and used as an antigen or a hapten, a specific antibody is obtained after the animal is immunized, and then the antibody is used for detecting the same kind of antigen substance in the tissue or cell. Since the complex of antigen and antibody is colorless, the site of antigen-antibody binding must also be visualized by histochemical methods to allow qualitative, localized or quantitative investigation of unknown antigens in tissues or cells.

In recent years, with the development of immunohistochemical techniques and the emergence of various specific antibodies, many difficult tumors have been clearly diagnosed. In routine pathological diagnosis of tumors, definitive morphological diagnosis is difficult to make in 5% -10% of cases by h.e. staining alone. In particular, the practical value of immunohistochemistry in tumor diagnosis and differential diagnosis is generally accepted, and the accuracy rate can reach 50 to 75 percent when the immunohistochemistry is used for differential diagnosis of low-differentiation or undifferentiated tumors.

Immunohistochemical experiments therefore represent a very large proportion of the in vitro diagnostic market. Carrying out conventional steps of immunohistochemical experiments, and preparing samples; antigen retrieval; permeating; eliminating endogenous peroxidase; sealing; primary antibody incubation; incubating and developing by using a secondary antibody; and (5) performing hematoxylin counterstaining. The duration of immunohistochemistry is about 3 hours, and shortening the immunohistochemistry time is very necessary for improving the in vitro diagnosis efficiency. Therefore, the invention effectively shortens the duration of immunohistochemical experiments.

Disclosure of Invention

The invention aims to provide a novel immunohistochemical three-in-one reagent formula to solve the technical problem of shortening immunohistochemical experiment time under the condition of not influencing experiment results.

In order to realize the purpose, the specific technical scheme of the novel immunohistochemical three-in-one reagent formula is as follows:

the utilization is transparent; eliminating endogenous peroxidase; the time of the steps such as sealing and the like is almost the same, corresponding reagents are prepared, and the three steps are integrated, so that the time of the immunohistochemical experiment is shortened;

the invention adopts the steps of preparing effective components with transparent immunohistochemistry, endogenous peroxidase elimination, sealing and the like into the same reagent without influencing the immunohistochemical experiment effect, and the specific preparation process is as follows:

transparent Triton X-100 (poly ethanol octyl phenyl ether), hydrogen peroxide for eliminating endogenous peroxidase, blocked goat serum, 0.2g 1% BSA, etc., and other components Tween-20, EDTA, PC300 dissolved in PBS (pH 7.4);

the optimal scheme is as follows: 0.1-0.4g Triton X-100; 1ml-4ml hydrogen peroxide; 0.5ml-2ml goat serum; 0.1g-0.4g BSA (bovine serum albumin); 0.01g to 0.04g of tween-20; 0.001g-0.01g EDTA; 0.1g-0.4g of PC300 and the like were dissolved in 17ml of PBS (pH 7.4).

Further, the feed additive comprises the following raw materials in parts by mass:

0.2g Triton X-100; 2ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.

Further, the feed additive comprises the following raw materials in parts by mass:

0.4g Triton X-100; 2ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.

Further, the feed additive comprises the following raw materials in parts by mass:

0.1g Triton X-100; 2ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.

Further, the feed additive comprises the following raw materials in parts by mass:

0.2g Triton X-100; 4ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.

Further, the feed additive comprises the following raw materials in parts by mass:

0.2g Triton X-100; 6ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.

Further, the feed additive comprises the following raw materials in parts by mass:

0.2g Triton X-100; 2ml of hydrogen peroxide; 10ml goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.

The novel immunohistochemical three-in-one reagent formula disclosed by the invention has the following advantages: the invention effectively shortens the immunohistochemical time and does not influence the immunohistochemical experiment effect.

Detailed Description

In order to better understand the purpose, structure and function of the present invention, the following will describe the formulation of a novel immunohistochemical three-in-one reagent of the present invention in detail with reference to the examples.

Example 1:

mixing 0.2g Triton X-100; 2ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g of PC300 and the like were dissolved in 17ml of PBS (pH 7.4);

example 2:

mixing 0.4g Triton X-100; 2ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g of PC300 and the like were dissolved in 17ml of PBS (pH 7.4).

Example 3:

mixing 0.1g Triton X-100; 2ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g of PC300 and the like were dissolved in 17ml of PBS (pH 7.4).

Example 4:

mixing 0.2g Triton X-100; 4ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g of PC300 and the like were dissolved in 17ml of PBS (pH 7.4).

Example 5:

mixing 0.2g Triton X-100; 6ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g of PC300 and the like were dissolved in 17ml of PBS (pH 7.4).

Example 6:

mixing 0.2g Triton X-100; 2ml of hydrogen peroxide; 10ml goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g of PC300 and the like were dissolved in 17ml of PBS (pH 7.4).

The test method is as follows:

the prepared immunohistochemical three-in-one reagent with components in different proportions is applied to immunohistochemical experiments, and the experimental effects of different formulas and the effect consistency rate when the three steps are respectively acted are observed.

The test results are shown in table 1, and the test results using the conventional experiment are shown in table 2:

table 1.

Sample (I)	Consistent rate of immunohistochemical experiment effect
		Example 1	100％
Example 2	98％
		Example 3	85％
Example 4	90％
		Example 5	92％
Example 6	90％

Table 2.

Sample (I)	Coloring ratio	Experiment time
			General experiments	98％±2％	185.5min

As can be seen from tables 1 and 2, the effect of embodiment 1 is the highest in agreement with the step-by-step experiment effect.

It is to be understood that the present invention has been described with reference to certain embodiments, and that various changes in the features and embodiments, or equivalent substitutions may be made therein by those skilled in the art without departing from the spirit and scope of the invention. In addition, many modifications may be made to adapt a particular situation or material to the teachings of the invention without departing from the essential scope thereof. Therefore, it is intended that the invention not be limited to the particular embodiment disclosed, but that the invention will include all embodiments falling within the scope of the appended claims.

Claims (7)

1. The novel immunohistochemical three-in-one reagent formula is characterized by comprising the following raw materials in parts by mass:

0.1-0.4g Triton X-100; 1ml-4ml hydrogen peroxide; 0.5ml-2ml goat serum; 0.1g-0.4g BSA; 0.01g to 0.04g of tween-20; 0.001g-0.01g EDTA; 0.1g-0.4g PC300 was dissolved in 17ml PBS, pH 7.4.

2. The novel immunohistochemical three-in-one reagent formula of claim 1, which is characterized by comprising the following raw materials in mass number:

0.2g Triton X-100; 2ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.

3. The novel immunohistochemical three-in-one reagent formula of claim 1, which is characterized by comprising the following raw materials in mass number:

0.4g Triton X-100; 2ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.

4. The novel immunohistochemical three-in-one reagent formula of claim 1, which is characterized by comprising the following raw materials in mass number:

0.1g Triton X-100; 2ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.

5. The novel immunohistochemical three-in-one reagent formula of claim 1, which is characterized by comprising the following raw materials in mass number:

0.2g Triton X-100; 4ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.

6. The novel immunohistochemical three-in-one reagent formula of claim 1, which is characterized by comprising the following raw materials in mass number:

0.2g Triton X-100; 6ml of hydrogen peroxide; 1ml of goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.

7. The novel immunohistochemical three-in-one reagent formula of claim 1, which is characterized by comprising the following raw materials in mass number:

0.2g Triton X-100; 2ml of hydrogen peroxide; 10ml goat serum; 0.2g 1% BSA; 0.02g Tween-20; 0.0067g EDTA; 0.2g PC300 in 17ml PBS, pH 7.4.