CN108359620B - Gram chromatin control tablet and its making method and use - Google Patents
Gram chromatin control tablet and its making method and use Download PDFInfo
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Abstract
The invention discloses a gram chromatin control tablet and a manufacturing method and application thereof, belongs to the technical field of staining, and solves the problem that the phenomena of false positive and false negative are easy to occur due to the fact that the color of a staining result cannot be accurately judged when bacteria are subjected to gram staining clinically and experimentally in the prior art. The invention provides a gram chromatin control wafer and a manufacturing method and application thereof, wherein the gram chromatin control wafer comprises a glass slide, a quality control bacterium area, a sample bacterium area and a protective cover, and the quality control wafer can effectively avoid the phenomena of false positive and false negative when the gram stain is carried out on bacteria clinically and experimentally and ensure the correctness of gram stain results.
Description
Technical Field
The invention belongs to the technical field of dyeing, and particularly relates to a gram chromatin control tablet and a manufacturing method and application thereof.
Background
In the prior art, the phenomenon of 'false positive' and 'false negative' easily occurs when gram staining is carried out on bacteria clinically and experimentally, generally caused by the quality of gram staining reagents or improper treatment time in staining, for example, the phenomenon of 'false negative' easily occurs when the decolorizing time is too long, and the phenomenon of 'false positive' easily occurs when the decolorizing time is too short. Clinically, antibiotics used for treating gram-positive bacteria and gram-negative bacteria are different, so the correctness of the staining result has a great influence on the reasonable prescription of the medicine by doctors.
The following methods are generally used in the prior art to ensure the correctness of gram staining experiments: (1) simply dyeing the sample bacteria (processing with dyes such as methylene blue, crystal violet or alkaline red), and microscopically examining the form of the sample bacteria, wherein the form of the sample bacteria can be spherical, rod-shaped, spiral or other special forms, but only one form; (2) if the sample bacteria are spherical after microscopic examination, culturing escherichia coli in a laboratory; if the sample bacteria are rod-shaped or spiral after microscopic examination, culturing staphylococcus aureus in a laboratory; (3) if the sample bacteria are spherical, performing mixed smear, drying, fixing and gram staining on the sample bacteria and escherichia coli at the same position of a glass slide until microscopic examination is performed, firstly observing whether the escherichia coli in a mixed smear area is red or not during the microscopic examination, indicating that the gram staining result is correct if the escherichia coli in the mixed smear area is red, and then observing the sample bacteria in the mixed smear area, wherein the sample bacteria are red, namely negative, and the sample bacteria are purple or bluish purple, namely positive; (4) if the sample bacteria are rod-shaped, the sample bacteria need to be subjected to mixed smear with staphylococcus aureus; (5) the mixed smear zone contains both the sample bacteria and Escherichia coli (or Staphylococcus aureus). However, the conventional method is time-consuming, and the sample bacteria need to be simply stained and subjected to microscopic examination each time, and the cultivation of escherichia coli or staphylococcus aureus can be determined after the morphology is known.
Meanwhile, the traditional gram staining method mainly adopts four-step methods of primary staining, mordant staining, decoloring and counterstaining, and most of students carry out optimization adjustment on the staining process of the traditional gram staining method so far, such as optimization screening on staining time and decoloring time or improvement on staining reagents; however, the above methods all require a large number of comparison references to achieve accurate staining results, and for the staining identification of large quantities of unknown bacteria, the method has low efficiency and high error rate, and in order to ensure the correctness of the staining results, each group of unknown bacteria needs to culture model strains of gram-positive bacteria and gram-negative bacteria, thereby increasing the workload.
Disclosure of Invention
1. Problems to be solved
The invention provides a gram chromatin control tablet and a manufacturing method and application thereof, aiming at the problem that the phenomena of false positive and false negative are easy to appear due to the fact that the color of a staining result cannot be accurately judged when the gram staining is carried out on bacteria in the prior art clinically and experimentally. The method can effectively avoid the phenomena of 'false positive' and 'false negative' when the bacteria are subjected to gram staining clinically and experimentally, and ensure the correctness of gram staining results.
2. Technical scheme
In order to solve the above problems, the present invention adopts the following technical solutions.
A method for preparing a gram chromatin control tablet comprises the following steps:
(1) preparing a glass slide, wherein one end of the glass slide is a marking area for information of a sample strain to be detected;
(2) the glass slide is provided with the quality control bacterium area and the sample bacterium area, the quality control bacterium area, the sample bacterium area and the marking area are not overlapped, and meanwhile, the number of the sample bacterium areas can be multiple, so that the treatment of multiple sample bacteria on the same glass slide is realized, and the efficiency is improved;
(3) preparing a waterproof adhesive tape and a double-sided waterproof rubber sheet, and bonding the waterproof adhesive tape and the double-sided waterproof rubber sheet together; then, perforating the waterproof adhesive tape and the double-sided waterproof adhesive sheet in a penetrating manner, then attaching one side of the waterproof adhesive tape to a quality control bacterium area to form a raised edge, and finally attaching the waterproof sticker to one side of the double-sided waterproof adhesive sheet to obtain the protective cover after ultraviolet irradiation treatment of the quality control bacterium area;
(4) respectively culturing escherichia coli and staphylococcus aureus; mixing the cultured escherichia coli liquid and staphylococcus aureus liquid to obtain a quality control liquid; then adding a quality control bacterial solution into the holes of the double-sided waterproof film of the protective cover to prepare a gram chromatin control film to be treated; the method of preparing the quality control bacterial liquid by mixing is convenient for observation under the same microscope visual field during microscopic examination after dyeing;
(5) and (3) drying the gram chromatin control wafer to be processed prepared in the step (4) until the gram chromatin control wafer to be processed can not see water traces, fixing the back surface of the gram chromatin control wafer to be processed, which is opposite to the quality control bacterium area, above the flame of an alcohol lamp for 2-3 times, then placing the hole of the double-sided waterproof film on the quality control bacterium area under an ultraviolet lamp with the wavelength of 260nm and irradiating the ultraviolet light for 15-25 min at a position of 25-30 cm under the ultraviolet lamp, preparing a waterproof sticker after irradiation processing, and attaching the waterproof sticker to the other side of the double-sided waterproof film. The gram chromatin control wafer to be treated is subjected to ultraviolet irradiation, so that other bacteria in the holes of the protective cover are completely killed, and the long-term storage of the quality control wafer is facilitated.
Preferably, the escherichia coli is escherichia coli strain CCTCC AB 93154; the staphylococcus aureus is staphylococcus aureus strain CCTCC AB 91093. Both of the above-mentioned strains (E.coli and S.aureus) were microbial type strains (http:// cctcc. whu. edu. cn /) of the China center for type culture Collection, and were not strains newly submitted by the inventors.
Preferably, the escherichia coli liquid is a culture liquid which controls the OD value of the cultivated escherichia coli to be 0.2-0.8; the staphylococcus aureus liquid is a culture liquid which controls the OD value of the cultivated staphylococcus aureus to be 0.2-0.8. The concentration of the bacteria is too large or too small, which has a certain influence on microscopic observation, for example, the bacteria are piled up in a microscopic field due to too large concentration, and then the bacteria are formed into a lump and cannot be distinguished in microscopic observation, and if the concentration of the bacteria is too small, escherichia coli and staphylococcus aureus cannot be observed simultaneously in the corresponding microscopic field.
Preferably, the quality control bacterial liquid is prepared by mixing the escherichia coli bacterial liquid and the staphylococcus aureus bacterial liquid according to the volume ratio of 5: 3. The mixing is carried out under the volume proportion, the escherichia coli and the staphylococcus aureus can reach the mixing ratio, so that the escherichia coli and the staphylococcus aureus in the microscopic visual field are uniformly distributed, because the proportion of the bacillus and the staphylococcus aureus is different, the discrimination of the microscopic visual field can be improved by proper proportion control, and the recognition in the shortest time can be realized.
Preferably, the drying temperature is controlled to be 35-45 ℃, and the drying treatment adopts an electrothermal blowing drying oven. Drying the quality control bacteria at the drying temperature within the range; meanwhile, the electrothermal drying treatment avoids the influence of water vapor caused by a wet-heat method.
Preferably, the drying temperature is controlled at 40 ℃, and the ultraviolet irradiation treatment time is 20 min. The drying temperature is selected to be 40 ℃, and the laminating tightness of escherichia coli and staphylococcus aureus with the glass slide is optimal; meanwhile, the ultraviolet irradiation treatment time is controlled to be 20min, and the inactivation effect of the thalli is optimal.
Preferably, the size of the aperture of the perforation in the step (A) is 5mm-6 mm; the thickness of the double-sided waterproof film is 3mm-6 mm. The hole diameter size of the punched hole and the thickness of the double-sided waterproof film are controlled within a proper range, and the probability of mixing of sample bacteria and quality control bacteria when the size ratio is too large or too small is further avoided;
the gram chromatin control tablet is manufactured by adopting the manufacturing method of the gram chromatin control tablet.
The application of the gram chromatin control tablet comprises the following steps:
(1) transferring the cultured sample bacteria to be detected to obtain a sample bacteria liquid; the sample bacteria to be detected can adopt solid cultures (bacterial colonies or lawn) of the sample bacteria, and can also adopt liquid cultures of the sample bacteria;
(2) selecting the gram chromatin control plate, and registering the information of the sample bacteria in the marking area; the confusion caused by the large number of bacteria in the sample to be detected is prevented;
(3) dropwise adding the sample bacterial liquid to a sample bacterial area, then performing smear, drying and fixing treatment on the sample bacterial liquid, and tearing off a protective cover of the gram chromatin control plate along the raised edge; simultaneously performing gram staining operation on a quality control bacterium area and a sample bacterium area in the gram chromatin control tablet, and dripping gram staining reagents to simultaneously cover the quality control bacterium and the sample bacterium with each staining reagent so as to achieve the purpose of simultaneous staining;
(4) performing microscopic examination after gram staining operation, firstly observing a staining area of the quality control bacteria, if red bacilli and purple cocci or bluish violet cocci are seen, indicating that the staining process is correct, and then observing the staining area of the sample bacteria, wherein purple or bluish violet indicates that the sample bacteria are gram-positive bacteria, and red indicates that the sample bacteria are gram-negative bacteria; if the staining area of the quality control bacteria is observed, the red bacillus, the purple coccus or the blue purple coccus is not seen, which indicates that the staining process fails, and the staining needs to be carried out again or the proper gram staining reagent needs to be replaced.
Preferably, the removing mode in the step (1) can be operated by using an inoculating loop.
3. Advantageous effects
Compared with the prior art, the invention has the beneficial effects that:
(1) the invention provides a method for manufacturing a gram chromatin control wafer, which adopts a structure of a protective cover, can protect quality control bacteria, prevent the quality control bacteria from being accidentally damaged, realize the simultaneous observation of the quality control bacteria and sample bacteria on the same glass slide, and improve the accuracy of gram staining results; meanwhile, a plurality of sample bacteria areas can be arranged, so that the detection of a plurality of sample bacteria on the same glass slide is realized;
(2) the invention provides a method for preparing gram chromatin control tablets, which mixes escherichia coli and staphylococcus aureus in quality control bacteria to prepare quality control bacteria liquid, is convenient for observation under the same visual field of a microscope and can accurately judge the accuracy of a dyeing result;
(3) the invention provides a method for manufacturing a gram chromatin control tablet, which is characterized in that the gram chromatin control tablet dropwise added with a quality control bacterium liquid is subjected to ultraviolet sterilization treatment, so that the long-term storage of the gram chromatin control tablet is realized;
(4) the invention provides a method for preparing gram chromatin control tablets, which controls escherichia coli and staphylococcus aureus in quality control bacteria to be diluted at a specific OD value and mixed according to a proportion, and the general method comprises the steps of simply dyeing sample bacteria, determining the form of the sample bacteria by microscopic examination, then mixing the sample bacteria with corresponding gram-positive or gram-negative bacteria with different forms, and then carrying out gram dyeing so as to facilitate the observation of the dyed quality control bacteria;
(5) the invention provides a method for manufacturing a gram chromatin control wafer, which can ensure that quality control bacteria can be better attached to a glass slide by selecting the drying temperature, thereby improving the quality of the gram chromatin control wafer;
(6) the invention provides an application of a gram chromatin control chip, which dyes quality control bacteria and sample bacteria simultaneously, and can judge the correctness of a dyeing result only by microscopic examination of the dyeing effect of the quality control bacteria.
Drawings
FIG. 1 is a schematic structural diagram of a gram chromatin control tablet according to the present invention;
FIG. 2(A) is a microscopic pattern of a quality-controlling bacterium according to example 1 of the present invention;
FIG. 2(B) is a micrograph of a sample strain of example 1 of the present invention;
FIG. 3(A) is a microscopic pattern of quality-controlling bacteria of example 2 of the present invention;
FIG. 3(B) is a micrograph of a sample strain of example 2 of the present invention;
FIG. 4(A) is a microscopic pattern of quality-controlling bacteria according to example 3 of the present invention;
FIG. 4(B) is a micrograph of the bacteria of example 3 of the present invention.
In the figure: 1. a marking region; 2. a quality control bacterium area; 3. a sample bacteria area; 4. a glass slide; 5. waterproof rubberized fabric; 6. a double-sided waterproof film; 7. waterproof paster.
Detailed Description
The invention is further described with reference to specific examples.
As shown in fig. 1, a method for preparing a gram chromatin control tablet comprises the following steps:
(1) preparing a glass slide 4, wherein one end of the glass slide 4 is a marking area 1 for recording information of strains;
(2) a quality control bacterium area 2 and a sample bacterium area 3 are arranged on a glass slide 4, the quality control bacterium area 2, the sample bacterium area 3 and a mark area 1 are not overlapped with each other and are arranged on the same surface of the glass slide 4, wherein the distance between the quality control bacterium area 2 and the sample bacterium area 3 is 2 mm;
(3) preparing a waterproof adhesive tape 5 (the waterproof adhesive tape 5 is an electrical adhesive tape) and a double-sided waterproof adhesive sheet 6 (the double-sided waterproof adhesive sheet 6 is a 3M double-sided adhesive sheet, the length and width of the double-sided waterproof adhesive sheet 6 are 1.2mm, and the thickness of the double-sided waterproof adhesive sheet is 3mm (in specific application, 3.2mm, 3.4mm, 3.6mm, 3.8mm, 4.0mm, 4.2mm, 4.4mm, 4.6mm, 4.8mm, 5.0mm, 5.2mm, 5.4mm, 5.6mm, 5.8mm or 6.0mm may be further taken), bonding the waterproof adhesive sheet 5 and the double-sided waterproof adhesive sheet 6 together, perforating the waterproof adhesive sheet 5 and the double-sided waterproof adhesive sheet 6 through a perforation, wherein the perforation has a pore size of 6mm (in specific application, 5.0mm, 5.1mm, 5.2mm, 5.3mm, 5.4mm, 5.5.5 mm, 5.5mm, 5.6mm, 5.5mm, 5mm, 5.5mm, 5.0mm, 5mm, 5.5mm, 5 The edge is convenient for tearing off the whole protective cover by hands at the later stage), the quality control bacterium area 2 is subjected to ultraviolet irradiation treatment, and finally the waterproof sticker 7 is attached to one surface of the double-sided waterproof film 6 to obtain the protective cover; the prior art needs to simply stain the sample bacteria each time for microscopic examination, and can determine to culture escherichia coli or staphylococcus aureus after knowing what form, but when the dyeing identification of mass unknown bacteria is faced, the simple dyeing and microscopic examination of the sample bacteria are required to be repeatedly carried out, then the mixture is mixed with escherichia coli or staphylococcus aureus for identification, the process is time-consuming and labor-consuming, and the quality control bacteria (escherichia coli or staphylococcus aureus) are required to be cultured in advance each time, the structure of the protective cover is innovatively designed on the basis of the glass slide, so that the quality control bacteria can be protected, meanwhile, the simultaneous observation of quality control bacteria and sample bacteria on the same glass slide can be realized (the steps of simple staining and microscopic examination of the sample bacteria are omitted), and the working efficiency is improved (the gram chromatin control wafer can be manufactured in advance in large batch);
(4) respectively culturing escherichia coli and staphylococcus aureus, wherein the escherichia coli is an escherichia coli strain CCTCC AB 93154, the staphylococcus aureus is a staphylococcus aureus strain CCTCC AB 91093, the escherichia coli and the staphylococcus aureus are cultured, the escherichia coli liquid is prepared by controlling the OD value of the cultured escherichia coli to be 0.2-0.8 (the OD value of the liquid of the cultured escherichia coli is generally within the range of 0.2-0.8, and the liquid of the cultured staphylococcus aureus does not need to be diluted), and the staphylococcus aureus liquid is prepared by controlling the OD value of the cultured staphylococcus aureus to be 0.2-0.8 (when the OD value of the liquid of the cultured staphylococcus aureus is generally more than 0.8, the liquid of the cultured staphylococcus aureus needs to be diluted); the escherichia coli bacterial fluid was then mixed with sterile normal saline in a ratio of 1: 19, diluting the staphylococcus aureus solution by 2 times, and mixing with sterile physiological saline in a ratio of 1: 49, mixing the escherichia coli liquid and the staphylococcus aureus liquid, and specifically mixing the escherichia coli liquid and the staphylococcus aureus liquid according to a volume ratio of 5:3 to obtain a quality control liquid; then adding a quality control bacterial solution into the holes of the double-sided waterproof film 6 on the protective cover to prepare a gram chromatin control film to be treated;
(5) drying the gram chromatin control wafer to be processed prepared in the step (4), wherein the drying temperature is controlled at 40 ℃ (when the drying temperature is specifically applied, 35 ℃, 36 ℃, 37 ℃, 38 ℃, 39 ℃, 41 ℃, 42 ℃, 43 ℃, 44 ℃ or 45 ℃), the selection of the drying temperature range leads the quality control bacteria to be better attached to the glass slide, when the temperature is too high (more than 70 ℃) or too low (less than 30 ℃), the adhesion force between the bacteria (the quality control bacteria) and the surface of the glass slide is insufficient, at the moment, the quality control bacteria are easy to fall off from the glass slide, further the dyeing of the quality control bacteria can not be realized, the drying process adopts an electrothermal blowing drying box until the gram chromatin control wafer to be processed can not see water stains, and then the back of the chromatin in the gram control wafer to be processed, which is opposite to the quality control bacteria area 2, is fixed above the flame of an alcohol lamp for 2 times, then, placing the hole of the double-sided waterproof film 6 on the quality control bacterium area 2 under an ultraviolet lamp with the wavelength of 260nm and irradiating for 15min by ultraviolet at a position of 25cm under the ultraviolet lamp, so that the long-term storage of the gram chromatin control chip is realized, and thus, when the mass of unknown bacteria are subjected to dyeing identification, only the gram chromatin control chip needs to be taken out, and the quality control bacteria do not need to be pre-cultured; after the irradiation treatment, the waterproof seal paper 7 is prepared and the waterproof seal paper 7 is attached to the other surface of the double-sided waterproof film 6.
The gram chromatin control tablet is manufactured by adopting the manufacturing method of the gram chromatin control tablet.
The application of gram chromatin control tablet includes the following steps
(1) Transferring the cultured sample bacteria to be detected to obtain a sample bacteria liquid; wherein the sample bacteria to be detected can also be lawn or colony, and is picked by using an inoculating loop;
(2) registering information of the sample bacteria in the marking area 1;
(3) selecting the gram chromatin control wafer, simultaneously dropwise adding the sample bacterial liquid to the sample bacterial area 3, then performing smear, drying and fixing treatment on the sample bacterial liquid, and tearing off the protective cover of the gram chromatin control wafer along the raised edge; simultaneously performing gram staining operation on a quality control bacterium area 2 and a sample bacterium area 3 in the gram chromatin control tablet, and performing microscopic examination after the gram staining operation;
(4) firstly, observing a staining area of quality control bacteria, if red bacilli and purple (bluish purple) cocci are seen, indicating that a staining result is correct, and then observing the staining area of sample bacteria, wherein purple (bluish purple) represents gram-positive bacteria, and red represents gram-negative bacteria; if the staining area of the quality control bacteria is observed, and the rhodobacter and the purple (bluish purple) coccus are not seen, the staining fails, and the staining needs to be carried out again or the proper gram staining reagent needs to be replaced.
In summary, in the prior art, in order to identify the staining of a large amount of unknown bacteria, the sample bacteria is usually stained to observe the morphology, then escherichia coli or staphylococcus aureus is cultured according to the shape, and finally whether the staining process of the sample bacteria has problems or not is judged by the microscopic examination of a mixed smear; this application creatively develops a gram chromatin control piece, specifically realize the protection to the quality control fungus in the quality control fungus district 2 through setting up the safety cover, this can once only make a large amount of gram chromatin control pieces, then when dyeing the identification to the sample fungus that awaits measuring, only need take out gram chromatin control piece and stain simultaneously to quality control fungus and sample fungus, if observe right dyeing result behind the dyeing of quality control fungus then explain the dyeing process failure, because the judgement that can accelerate the dyeing result has been made in advance to the quality control fungus, this is unobvious to technical staff in the field.
Example 1
The gram chromatin control tablet of the present example was used as follows,
(1) transferring the cultured sample bacteria to be detected by using an inoculating loop to obtain sample bacteria liquid;
(2) registering information of the sample bacteria in the labeling area 1, wherein the information is bacillus subtilis CCTCC AB90008, gram-positive bacteria;
(3) selecting a gram chromatin control plate, simultaneously dropwise adding a sample bacterial liquid to the sample bacterial area 3, then performing smear, drying and fixing treatment on the sample bacterial liquid, and tearing off a protective cover of the gram chromatin control plate along the raised edge; simultaneously performing gram staining operation on a quality control bacterium area 2 and a sample bacterium area 3 in the gram chromatin control tablet, and performing microscopic examination after the gram staining operation;
(4) first, the staining area of the quality control bacteria is observed, so that rhodobacter and purple (bluish purple) coccus are seen, the microscopic pattern of the quality control bacteria is shown in fig. 2(a), which shows that the staining result is correct, and then the staining area of the sample bacteria is observed, wherein purple (bluish purple) represents gram-positive bacteria, and the microscopic pattern of the sample bacteria is shown in fig. 2 (B).
Example 2
The gram chromatin control tablet of the present example was used as follows,
(1) transferring the cultured sample bacteria to be detected by using an inoculating loop to obtain sample bacteria liquid;
(2) registering information on the sample bacteria in the labeling zone 1, such as proteus CCTCC AB 91103, gram-negative bacteria;
(3) selecting a gram chromatin control plate, simultaneously dropwise adding a sample bacterial liquid to the sample bacterial area 3, then performing smear, drying and fixing treatment on the sample bacterial liquid, and tearing off a protective cover of the gram chromatin control plate along the raised edge; simultaneously performing gram staining operation on a quality control bacterium area 2 and a sample bacterium area 3 in the gram chromatin control tablet, and performing microscopic examination after the gram staining operation;
(4) first, the staining area of the quality control bacteria is observed, so that rhodobacter and purple (bluish purple) coccus are seen, the microscopic pattern of the quality control bacteria is shown in fig. 3(a), which indicates that the staining result is correct, and then the staining area of the sample bacteria is observed, wherein red indicates gram-negative bacteria, and the microscopic pattern of the sample bacteria is shown in fig. 3 (B).
Example 3
The gram chromatin control tablet of the present example was used as follows,
(1) transferring the cultured sample bacteria to be detected by using an inoculating loop to obtain sample bacteria liquid;
(2) registering information on the sample bacteria in marker zone 1, as follows, Micrococcus luteus CMCC (B)28001, gram positive bacteria;
(3) selecting a gram chromatin control plate, simultaneously dropwise adding a sample bacterial liquid to the sample bacterial area 3, then performing smear, drying and fixing treatment on the sample bacterial liquid, and tearing off a protective cover of the gram chromatin control plate along the raised edge; simultaneously performing gram staining operation on a quality control bacterium area 2 and a sample bacterium area 3 in the gram chromatin control tablet, and performing microscopic examination after the gram staining operation;
(4) first, the staining area of the quality control bacteria is observed, so that rhodobacter and purple (bluish purple) coccus are seen, the microscopic pattern of the quality control bacteria is shown in fig. 4(A), which shows that the staining result is correct, and then the staining area of the sample bacteria is observed, wherein purple (bluish purple) represents gram-positive bacteria, and the microscopic pattern of the sample bacteria is shown in fig. 4 (B).
Claims (9)
1. A method for preparing a gram chromatin control tablet is characterized by comprising the following steps:
(1) preparing a glass slide (4), wherein one end of the glass slide (4) is a marking area (1) for recording the information of the strain of the sample to be detected;
(2) a quality control bacterium area (2) and a sample bacterium area (3) are arranged on the glass slide (4), and the quality control bacterium area (2), the sample bacterium area (3) and the marking area (1) are not overlapped with each other;
(3) preparing a waterproof adhesive tape (5) and a double-sided waterproof adhesive sheet (6), wherein the thickness of the double-sided waterproof adhesive sheet (6) is 3-6 mm, and bonding the waterproof adhesive tape (5) and the double-sided waterproof adhesive sheet (6) together; then, the waterproof adhesive tape (5) and the double-sided waterproof adhesive sheet (6) are perforated, and one side of the waterproof adhesive tape (5) is attached to the quality control bacterium area (2) to form a raised edge; after the quality control bacterium area (2) is subjected to ultraviolet irradiation treatment, finally, a waterproof sticker (7) is attached to one surface of the double-sided waterproof film (6); manufacturing a protective cover;
(4) respectively culturing escherichia coli and staphylococcus aureus; mixing the cultured escherichia coli liquid and staphylococcus aureus liquid to obtain a quality control liquid; then adding a quality control bacterial solution into the holes on the double-sided waterproof film (6) of the protective cover to prepare a gram chromatin control film to be processed;
(5) drying the gram chromatin control wafer to be processed prepared in the step (4), fixing the back surface of the gram chromatin control wafer to be processed, which is opposite to the quality control bacterium area (2), above the flame of an alcohol lamp for 2-3 times, placing the hole on the double-sided waterproof film (6) of the quality control bacterium area (2) under an ultraviolet lamp with the wavelength of 260nm at the position of 25cm-30cm, irradiating by ultraviolet for 15min-25min, preparing a waterproof sticker (7) after irradiation treatment, and attaching the waterproof sticker (7) to the other side of the double-sided waterproof film (6).
2. The method of claim 1, wherein the method comprises: the Escherichia coli liquid is a culture liquid which controls the OD value of the cultured Escherichia coli to be 0.2-0.8; the staphylococcus aureus liquid is a culture liquid which controls the OD value of the cultivated staphylococcus aureus to be 0.2-0.8.
3. A method for preparing a gram chromatin control tablet according to claim 2, wherein: the quality control bacterial liquid is prepared by mixing the escherichia coli bacterial liquid and the staphylococcus aureus bacterial liquid according to the volume ratio of 5: 3.
4. The method of claim 1, wherein the method comprises: the temperature of the drying treatment in the step (5) is controlled to be 35-45 ℃, and the drying treatment adopts an electrothermal blowing drying oven.
5. A method for preparing a gram chromatin control tablet according to claim 3, wherein: the temperature of the drying treatment in the step (5) is controlled at 40 ℃, and the ultraviolet irradiation treatment time is 20 min.
6. The method of claim 1, wherein the method comprises: the aperture size of the punching in the step (3) is 5mm-6 mm.
7. A gram chromatin control tablet, comprising: the gram chromatin control tablet of claim 1, which is prepared by the method.
8. The application of a gram chromatin control tablet is characterized in that: the method comprises the following steps:
(1) transferring the cultured sample bacteria to be detected to obtain a sample bacteria liquid;
(2) a gram chromatin control chip as claimed in claim 7 wherein information on the sample bacteria is registered at the label zone (1);
(3) dropwise adding the sample bacterial liquid to the sample bacterial area (3), then performing smear, drying and fixing treatment on the sample bacterial liquid, and tearing off the protective cover of the gram chromatin control plate along the raised edge; simultaneously performing gram staining operation on a quality control bacterium area (2) and a sample bacterium area (3) in the gram chromatin control tablet;
(4) performing microscopic examination after gram staining operation, firstly observing a staining area of the quality control bacteria, if red bacilli and purple cocci or bluish violet cocci are seen, indicating that the staining process is correct, and then observing the staining area of the sample bacteria, wherein purple or bluish violet indicates that the sample bacteria are gram-positive bacteria, and red indicates that the sample bacteria are gram-negative bacteria; if the staining area of the quality control bacteria is observed, the red bacillus, the purple coccus or the blue purple coccus is not seen, which indicates that the staining process fails, and the staining needs to be carried out again or the proper gram staining reagent needs to be replaced.
9. Use of a gram chromatin control tablet according to claim 8, wherein: in the step (1), the moving mode is to operate by adopting an inoculating loop.
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