Kit for detecting typhoid and paratyphoid and application thereof
阅读说明：本技术 检测伤寒与副伤寒的试剂盒及其应用 (Kit for detecting typhoid and paratyphoid and application thereof ) 是由 张小龙 于 2019-09-25 设计创作，主要内容包括：本发明提供了一种检测伤寒与副伤寒的试剂盒,包括：V型底的微量反应板、沙黄溶液、肥达氏抗原和外斐氏抗原；所述肥达氏抗原包括：伤寒沙门氏菌O、伤寒沙门氏菌H、甲型副伤寒沙门氏菌、乙型副伤寒沙门氏菌和丙型副伤寒沙门氏菌；所述外斐氏抗原包括：变形杆菌OX19、变形杆菌OX2和变形杆菌OXK。在检测过程中,阳性血清样本与所述肥达氏抗原或所述外斐氏抗原在微量反应板孔底的形成的凝集,配合以沙黄溶液与所述肥达氏抗原和外斐氏抗原的显色作用,使得对于伤寒与副伤寒的检测结果判断更清晰、准确。(The invention provides a kit for detecting typhoid and paratyphoid, which comprises: a V-shaped bottom micro-reaction plate, a Shahuang solution, a Felidae antigen and a Murphy antigen; the widal antigen includes: salmonella typhi O, Salmonella typhi H, Salmonella paratyphi A, Salmonella paratyphi B, and Salmonella paratyphi C; the exofei antigen comprises: bacillus proteus OX19, Bacillus proteus OX2, and Bacillus proteus OXK. In the detection process, the positive serum sample and the feddal antigen or the ectofephne antigen are agglutinated at the bottom of the hole of the micro-reaction plate, and the chromogenic action of the saffron solution and the feddal antigen and the ectofephne antigen is matched, so that the detection result judgment of the typhoid fever and the paratyphoid fever is clearer and more accurate.)
1. A kit for detecting typhoid and paratyphoid is characterized by comprising:
a V-shaped bottom micro-reaction plate, a saffron reagent, a feudal antigen and a ectofei antigen;
the widal antigen includes: salmonella typhi O, Salmonella typhi H, Salmonella paratyphi A, Salmonella paratyphi B, and Salmonella paratyphi C;
the exofei antigen comprises: bacillus proteus OX19, Bacillus proteus OX2, and Bacillus proteus OXK.
2. The kit for detecting typhoid and paratyphoid as claimed in claim 1, wherein the V-bottom microplate is an optically transparent V-bottom 96-well plate.
3. The kit for detecting typhoid and paratyphoid as claimed in claim 1, wherein the saffron reagent is a saffron solution, and the concentration of the saffron solution is 0.3-0.7%.
4. The kit for detecting typhoid and paratyphoid as claimed in claim 1, wherein the storage concentrations of widal antigens are as follows:
the concentration of the salmonella typhosa O is 6.0 multiplied by 109～8.0×109Bacterial count/mL;
the concentration of the salmonella typhosa H is 6.0 x 109～8.0×109Bacterial count/mL;
the concentration of the salmonella paratyphi A is6.0×109～8.0×109Bacterial count/mL;
the concentration of the salmonella paratyphi B is 6.0 multiplied by 109～8.0×109Bacterial count/mL;
the concentration of the salmonella paratyphi C is 6.0 multiplied by 109～8.0×109Bacterial count/mL;
the storage concentrations of the ectofei antigen were as follows:
the concentration of the proteus OX19 was 6.0X 109～8.0×109Bacterial count/mL;
the concentration of the proteus OX2 was 6.0X 109～8.0×109Bacterial count/mL;
the concentration of the proteus OXK is 6.0 x 109～8.0×109bacteria/mL.
5. The kit for detecting typhoid and paratyphoid as claimed in claim 4, wherein the storage temperatures of the feudal antigen and the exothermia antigen are: 2-8 ℃.
6. The kit for detecting typhoid and paratyphoid as claimed in any one of claims 1 to 5, wherein the concentrations of the widal antigens when the kit is used are as follows:
the concentration of the salmonella typhosa O is 6.0 multiplied by 108～8.0×108Bacterial count/mL;
the concentration of the salmonella typhosa H is 6.0 x 108～8.0×108Bacterial count/mL;
the concentration of the salmonella paratyphi A is 6.0 multiplied by 108～8.0×108Bacterial count/mL;
the concentration of the salmonella paratyphi B is 6.0 multiplied by 108～8.0×108Bacterial count/mL;
the concentration of the salmonella paratyphi C is 6.0 multiplied by 108～8.0×108Bacterial count/mL;
the concentrations of the ectofei antigen were as follows:
the concentration of the proteus OX19 is 6.0108～8.0×108Bacterial count/mL;
the concentration of the proteus OX2 was 6.0X 108～8.0×108Bacterial count/mL;
the concentration of the proteus OXK is 6.0 x 108～8.0×108bacteria/mL.
7. The kit for detecting typhoid and paratyphoid as claimed in claim 6, wherein the V-bottom microplate is divided into at least eight groups of wells at the time of use of the kit;
mixing the salmonella typhi O, the sallow solution and a first group of serum to be detected in a first group of holes of the V-shaped bottom micro reaction plate;
mixing the salmonella typhi H, the sallow solution and a second group of serum to be detected in a second group of holes of the V-shaped bottom micro reaction plate;
in the third group of holes of the V-shaped bottom micro reaction plate, the salmonella paratyphi A, the sallow solution and the third group of serum to be detected are mixed;
in a fourth group of holes of the V-shaped bottom micro reaction plate, the salmonella paratyphi B, the sallow solution and a fourth group of serum to be detected are mixed;
in a fifth group of holes of the V-shaped bottom micro reaction plate, the salmonella paratyphi C, the sallow solution and a fifth group of serum to be detected are mixed;
in a sixth group of holes of the V-shaped bottom microplate, the proteus OX19, the saffron solution and a sixth group of serum to be tested are mixed;
in a seventh group of holes of the V-shaped bottom microplate, the proteus OX2, the saffron solution and a seventh group of serum to be tested are mixed;
and in the eighth group of holes of the V-shaped bottom micro reaction plate, the proteus OXK, the saffron solution and the eighth group of serum to be detected are mixed.
8. The kit for detecting typhoid and paratyphoid as claimed in claim 7, wherein the volume ratio of the salmonella typhi O, the salmonella typhi H, the salmonella paratyphi A, the salmonella paratyphi B, the salmonella paratyphi C, the proteus OX19, the proteus OX2 and the proteus OXK to the safrole solution is 1: (9-11).
9. The kit for detecting typhoid and paratyphoid as claimed in any one of claims 1 to 5, 7 and 8, wherein the V-bottom microplate is incubated in an electrothermal incubator at 35-37 ℃ for 16-20 hours to interpret the result.
10. The use of the kit for detecting typhoid and paratyphoid as defined in any one of claims 1 to 9 for the preparation of an in vitro auxiliary diagnostic kit for detecting infectious diseases caused by typhoid and paratyphoid.
Technical Field
The invention belongs to the field of medical inspection, and particularly relates to a kit for detecting typhoid and paratyphoid and application thereof.
Background
Typhoid fever is an enteric infectious disease caused by a typhoid serotype in a salmonella enterica subspecies, and paratyphoid fever is a disease similar to typhoid fever caused by a paratyphoid A or B or C serotype in the salmonella enterica subspecies. The symptoms of paratyphoid A and paratyphoid B are similar to those of typhoid fever, but the general condition is mild, the course of disease is short, and the fatality rate is low; the side cold C symptoms are different and can be manifested as mild typhoid fever, acute gastroenteritis or sepsis. Typhoid and paratyphoid can be outbreak and epidemic due to water source and food pollution.
The fedah reaction and the exothermia are traditional methods for detecting typhoid fever and paratyphoid fever, but the traditional detection methods of the fedah reaction and the exothermia are mainly tube agglutination methods, which mainly have the following problems: 1. the operation is more complicated, a large number of test tubes are needed, and the sample adding amount of the required detection reagent is large. 2. The result is difficult to clearly judge by naked eyes after the water bath box is incubated, and whether the agglutination reaction completely occurs is difficult to judge.
Disclosure of Invention
Based on this, the invention aims to provide a kit for detecting typhoid and paratyphoid and application thereof, so that the method for detecting typhoid and paratyphoid is simpler and more convenient, the result is clearer, the consumption of detection reagents can be reduced, and the cost is saved.
The specific technical scheme is as follows:
a kit for detecting typhoid and paratyphoid, comprising:
a V-shaped bottom micro-reaction plate, a saffron reagent, a feudal antigen and a ectofei antigen;
the widal antigen includes: salmonella typhi O, Salmonella typhi H, Salmonella paratyphi A, Salmonella paratyphi B, and Salmonella paratyphi C;
the exofei antigen comprises: bacillus proteus OX19, Bacillus proteus OX2, and Bacillus proteus OXK.
The invention also provides application of the kit for the fedaofil test, and the specific technical scheme is as follows:
the kit for detecting typhoid and paratyphoid is applied to the preparation of an in-vitro auxiliary diagnostic reagent kit for detecting infectious diseases caused by typhoid and paratyphoid.
Based on the technical scheme, the invention has the following beneficial effects:
the kit for detecting the typhoid and paratyphoid comprises a V-type base micro reaction plate, a Feddler antigen, a Murphy's antigen and a safrole reagent, when the kit is used for detecting the typhoid and paratyphoid, each Feddler antigen and Murphy's antigen are respectively mixed with a safrole solution and then respectively mixed with a serum sample to be detected, wherein the serum possibly containing antibodies aiming at the typhoid and/or paratyphoid is subjected to agglutination reaction with salmonella typhi O, salmonella typhi H, salmonella paratyphoid A, salmonella paratyphoid B and salmonella paratyphoid C, and Proteus OX19, Proteus OX2 and Proteus OXK have a common surface antigen with rickettsia and are subjected to agglutination reaction with serum possibly containing antibodies aiming at the rickettsia, and during the detection process, the positive serum sample is agglutinated with the Feddler antigen or the ectofephus antigen formed on the micro reaction plate hole base, the chromogenic effect of the saffron solution and the feudal antigen and the exothermia antigen is matched, so that the detection results of the typhoid and paratyphoid are judged more clearly and accurately.
When being used for detecting typhoid and paratyphoid with this kit, for traditional feuda, the reaction of weifei adopts the test tube to test, when adopting the micro reaction plate at V type end, after hatching, the thallus that does not take place agglutination reaction with the antibody deposits in the reaction plate bottom along V type end more concentratedly, antigen-antibody takes place the agglutination reaction after, the quincunx type agglutination of production changes and distinguishes through the naked eye, and need not traditional feuda, need observe the clarity earlier when weifeihe reaction adopts the test tube to detect, rock the more complicated step of test tube observation gently again, complex operation nature in this step has been avoided, also avoided rocking the improper error that leads to observing the production of dynamics.
Moreover, the micro reaction plate with the V-shaped bottom is a normalized product generally, and is adaptive to the row-type sample adding gun, so that the micro reaction plate is beneficial to being matched with the row-type sample adding gun for detection, on one hand, the micro reaction plate only consumes a small amount of detection reagent in each detection, on the other hand, the row-type sample adding improves the detection efficiency, and meanwhile, the error caused by the multi-test tube fractional sample adding in the traditional method can be avoided as much as possible.
The kit provided by the invention is beneficial to improving the efficiency of in-vitro auxiliary diagnosis of infectious diseases caused by typhoid and paratyphoid, and is suitable for clinical popularization.
Drawings
FIGS. 1-2 are the results of serum tests using the kit of example 1 for detecting typhoid and paratyphoid;
FIG. 3 is a graph showing the results of the detection of positive control serum (++);
FIGS. 4-8 are exemplary graphs corresponding to the results of each agglutination reaction (++++), (+++), (+ -) -and (-);
FIG. 9 is a diagram showing the results of detection of negative control serum;
FIGS. 10-14 are schematic illustrations of different agglutination results detected using conventional methods;
FIG. 15 is a graph showing the results of the positive sera of comparative example 2;
FIG. 16 is a graph showing the results of the negative serum of comparative example 2;
FIG. 17 is a graph showing the results of the measurement in comparative example 3;
FIG. 18 is a graph showing the results of the positive sera of comparative example 4.
Detailed Description
In order that the invention may be more readily understood, reference will now be made to the following more particular description of the invention, examples of which are set forth below. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein. These embodiments are provided so that this disclosure will be thorough and complete. It is to be understood that the experimental procedures in the following examples, where specific conditions are not noted, are generally in accordance with conventional conditions, or with conditions recommended by the manufacturer. The various reagents used in the examples are commercially available.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. As used herein, the term "and/or" includes any and all combinations of one or more of the associated listed items.
A kit for detecting typhoid and paratyphoid is characterized by comprising:
a V-shaped bottom micro-reaction plate, a saffron reagent, a feudal antigen and a ectofei antigen;
the widal antigen includes: salmonella typhi O, Salmonella typhi H, Salmonella paratyphi A, Salmonella paratyphi B, and Salmonella paratyphi C;
the exofei antigen comprises: bacillus proteus OX19, Bacillus proteus OX2, and Bacillus proteus OXK.
Preferably, the V-bottom microplate is an optically transparent V-bottom 96-well plate.
Preferably, the safranine reagent is a safranine solution. More preferably, the concentration of the saffron solution is 0.3% to 0.7%.
Preferably, the storage concentrations of the feudal antigens are as follows:
the concentration of the salmonella typhosa O is 6.0 multiplied by 109～8.0×109Bacterial count/mL;
the concentration of the salmonella typhosa H is 6.0 x 109～8.0×109Bacterial count/mL;
the concentration of the salmonella paratyphi A is 6.0 multiplied by 109～8.0×109Bacterial count/mL;
the concentration of the salmonella paratyphi B is 6.0 multiplied by 109～8.0×109Bacterial count/mL;
the C-type paratyphoid salmonellaThe concentration of the bacteria is 6.0 × 109～8.0×109Bacterial count/mL;
the storage concentrations of the ectofei antigen were as follows:
the concentration of the proteus OX19 was 6.0X 109～8.0×109Bacterial count/mL;
the concentration of the proteus OX2 was 6.0X 109～8.0×109Bacterial count/mL;
the concentration of the proteus OXK is 6.0 x 109～8.0×109bacteria/mL.
Preferably, the storage temperature of the fedah antigen and the ectofei antigen is: 2-8 ℃.
Preferably, the concentrations of the feudal antigens when the kit is used are as follows:
the concentration of the salmonella typhosa O is 6.0 multiplied by 108～8.0×108Bacterial count/mL;
the concentration of the salmonella typhosa H is 6.0 x 108～8.0×108Bacterial count/mL;
the concentration of the salmonella paratyphi A is 6.0 multiplied by 108～8.0×108Bacterial count/mL;
the concentration of the salmonella paratyphi B is 6.0 multiplied by 108～8.0×108Bacterial count/mL;
the concentration of the salmonella paratyphi C is 6.0 multiplied by 108～8.0×108Bacterial count/mL;
the concentrations of the ectofei antigen were as follows:
the concentration of the proteus OX19 was 6.0X 108～8.0×108Bacterial count/mL;
the concentration of the proteus OX2 was 6.0X 108～8.0×108Bacterial count/mL;
the concentration of the proteus OXK is 6.0 x 108～8.0×108bacteria/mL.
Preferably, in use of the kit, the V-bottom microplate is divided into at least eight sets of wells;
mixing the salmonella typhi O, the sallow solution and a first group of serum to be detected in a first group of holes of the V-shaped bottom micro reaction plate;
mixing the salmonella typhi H, the sallow solution and a second group of serum to be detected in a second group of holes of the V-shaped bottom micro reaction plate;
in the third group of holes of the V-shaped bottom micro reaction plate, the salmonella paratyphi A, the sallow solution and the third group of serum to be detected are mixed;
in a fourth group of holes of the V-shaped bottom micro reaction plate, the salmonella paratyphi B, the sallow solution and a fourth group of serum to be detected are mixed;
in a fifth group of holes of the V-shaped bottom micro reaction plate, the salmonella paratyphi C, the sallow solution and a fifth group of serum to be detected are mixed;
in a sixth group of holes of the V-shaped bottom microplate, the proteus OX19, the saffron solution and a sixth group of serum to be tested are mixed;
in a seventh group of holes of the V-shaped bottom microplate, the proteus OX2, the saffron solution and a seventh group of serum to be tested are mixed;
and in the eighth group of holes of the V-shaped bottom micro reaction plate, the proteus OXK, the saffron solution and the eighth group of serum to be detected are mixed.
Preferably, the volume ratio of the salmonella typhi O, the salmonella typhi H, the salmonella paratyphi A, the salmonella paratyphi B, the salmonella paratyphi C, the proteus OX19, the proteus OX2 and the proteus OXK to the salsa yellow solution is 1: (9-11).
Preferably, the kit for detecting typhoid and paratyphoid further comprises sterile physiological saline.
Preferably, when the kit is used, the V-shaped bottom micro reaction plate is placed in an electrothermal constant temperature incubator at 35-37 ℃ for incubation for 16-20 hours, and then the result is interpreted.
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