A test piece for preventing liquefaction of colony count and a preparation method and application thereof

Abstract

The application discloses a kind of anti-liquefaction colony total number test pieces and its preparation method and application.A kind of anti-liquefaction colony total number test piece, the test piece includes transparent upper layer film and bottom plate arranged in each other, be equipped with anti-liquefaction cold water gel and anti-liquefaction additive on the transparent upper layer film, the bottom plate is equipped with the mixed powder of colony total number chromogenic culture medium and compound cold water gel.The application develops a kind of colony chromogenic clear and resistant spore class bacillus decomposition liquefaction, and can be quickly and accurately counted colony total number test piece is of great significance to the food rapid detection industry of our country.

Description

Technical Field

[0001] This invention relates to the field of colony testing technology, specifically to an anti-liquefaction total colony count test strip, its preparation method, and its application. Background Technology

[0002] Total bacterial count, as a crucial indicator of food hygiene, holds a particularly important position, being one of the indicator bacteria with the most mandatory testing items across various food categories. As of 2020, approximately 38 categories (types) of food in my country's product standards specify limits for total bacterial count. Among these, 36 categories (types) employ a three-level sampling scheme (n=5), resulting in a substantial testing volume. In final product testing and hygiene monitoring of the processing environment, in-process products, and raw materials, total bacterial count is renowned for its high testing frequency and large testing volume. With the development of the food industry, the requirements for testing methods are increasing, and methods such as the test strip method—simple, accurate, and environmentally friendly—are becoming increasingly well-known and accepted by laboratories. Therefore, GB 4789.2-2022, "National Food Safety Standard - Microbiological Examination of Food: Determination of Total Bacterial Count," added the test strip method for parallel access.

[0003] Microbial test strips eliminate the need for extensive auxiliary work such as preparing culture media, sterilizing, and cleaning culture vessels. They are convenient to use, highly accurate, and require only three simple steps: sample preparation, inoculation, and incubation to count microorganisms, significantly reducing testing time. The microbial disc method, developed in recent years, uses a breathable and water-resistant polymer material as a carrier and a cold water-soluble gel as a solidifying agent, making it a ready-to-use sterile product. For example, Chinese invention patent CN105754849B discloses a cold water gel test strip for total bacterial count that is simple to operate, inexpensive, and accurate in detection. The test strip consists of a transparent plastic upper film with an adhesive inner side, a plastic middle layer with a hollowed-out area, and a base plate printed with squares. The materials are heat-resistant to 130℃~150℃. The hollowed-out area of ​​the plastic middle layer is circular, elliptical, or square, and the ratio of the hollowed-out area to the middle layer area is 1:1.6~2.7. The hollowed-out area is filled with a colorimetric culture medium for total bacterial count detection. The inner side of the transparent plastic upper film is coated with a compound cold water soluble gel. The colorimetric culture medium for total bacterial count detection, in addition to the basal medium of tryptone, yeast extract, sodium chloride, and glucose, also includes the colorimetric reagent TTC (2,3,5-triphenyltetrazolium chloride), the growth factor sodium pyruvate, the surfactant Tween 80, mannitol, and the loosening agent soybean residue. The soybean residue loosening agent is prepared by drying fresh soybean residue in a 50-60℃ constant temperature drying oven, grinding it, and passing it through a 100-120 mesh sieve. The compounded cold water-soluble gel consists of guar gum, sodium polyacrylate, and sodium carboxymethyl cellulose. The weight composition of the colorimetric culture medium for total bacterial count detection is: 15-30g tryptone, 5-15g yeast extract, 5-10g sodium chloride, 1-5g glucose, 1-3g sodium pyruvate, 1-3g mannitol, 1-3mL Tween 80, 0.2-0.5g TTC, and 3-10g soybean residue. The compound cold water soluble gel has the following weight components: 8g guar gum, 1-2g sodium polyacrylate and 1-2g sodium carboxymethyl cellulose.

[0004] While these test strips offer significant advantages in terms of convenience and practicality, they still have some shortcomings as a new technology for microbial detection. For example, when food samples contain Bacteroides spores, they can rapidly grow and multiply within a short incubation period, producing large amounts of enzymes that cause most commercially available microbial test strips to liquefy, resulting in colonies that coalesce and become difficult to observe and count, thus reducing accuracy. Summary of the Invention

[0005] To address the shortcomings of existing technologies, the present invention aims to provide a liquefaction-resistant total bacterial count test strip, its preparation method, and its application. The present invention develops a total bacterial count test strip with clear colony color development and resistance to decomposition and liquefaction by Bacillus spores, which can be rapidly and accurately counted, and is of great significance to my country's rapid food testing industry.

[0006] The technical problem to be solved by the present invention is achieved through the following technical solution:

[0007] In a first aspect, there is a liquefaction-resistant total bacterial count test strip, the test strip comprising a transparent upper membrane and a base plate stacked on top of each other, wherein the transparent upper membrane is provided with a liquefaction-resistant cold hydrogel and an anti-liquefaction additive, and the base plate is provided with a mixture of a total bacterial count colorimetric culture medium and a compounded cold hydrogel powder.

[0008] In this invention, the liquefaction-resistant cold hydrogel is a high-acyl gelling gel; the anti-liquefaction additive is glycine, and the dosage is 2% to 10% of the cold hydrogel powder.

[0009] Secondly, a method for preparing the above-mentioned anti-liquefaction bacterial count test strip includes the following steps:

[0010] (1) Preparation: liquefaction-resistant cold hydrogel, anti-liquefaction additive, total bacterial count colorimetric culture medium and mixed powder of compound cold hydrogel;

[0011] (2) Spray the liquefaction-resistant cold hydrogel and anti-liquefaction additive on the inside of the transparent upper membrane; spray the mixed powder of the total colony count colorimetric culture medium and the compound cold hydrogel on the inside of the base plate;

[0012] The transparent upper layer film is laminated and bonded to the base plate;

[0013] (3) Sterilization.

[0014] The application of the anti-liquefaction total bacterial count test strip as described in claim 1 in detecting the total bacterial count in food.

[0015] Compared with the prior art, the present invention has the following beneficial effects:

[0016] This invention relates to an anti-liquefaction total bacterial count test strip. Because it uses a cold hydrogel that is not easily decomposed by spore-forming bacteria, and incorporates a special anti-liquefaction additive mixed with the cold hydrogel powder, samples containing spore-forming bacteria do not liquefy or diffuse during cultivation, resulting in clear, easily observable, and countable single colonies. Since both the base plate and the top membrane contain cold hydrogel, the sample solution automatically diffuses into a circular culture area upon addition. Furthermore, the gel rapidly solidifies after absorbing water, eliminating the need for external pressure to form a circular culture area, thus preventing sample leakage. The operation is simple, and the results are accurate.

[0017] Furthermore, this invention also selects growth-promoting factors sodium pyruvate and glycine, which have the effect of promoting and repairing bacterial growth. During the culture process, the bacteria achieve rapid growth, and the addition of the colorimetric reagent TTC makes the colonies appear red on the test strip, enabling successful counting in a short time. Attached Figure Description

[0018] Figure 1 A schematic diagram showing the performance of Bacillus subtilis on the test strip of this invention;

[0019] Figure 2 This is a schematic diagram showing the performance of Bacillus subtilis on the test strip of Comparative Example 1.

[0020] Figure 3 This is a schematic diagram showing the performance of Bacillus subtilis on the test strip of Comparative Example 2.

[0021] Figure 4 The correlation curves between the test piece method and the national standard method of this invention. Detailed Implementation

[0022] As described in the background section, existing test strips have the significant advantages of being convenient and practical, but as a new technology for microbial detection, they still have some shortcomings. For example, when food samples contain Bacillus spores, they can rapidly grow and multiply within a short culture time, producing a large amount of enzymes that cause most commercially available microbial test strips to liquefy, making the colonies difficult to observe and count due to their merging, thus reducing accuracy.

[0023] To address the aforementioned technical problems, this invention proposes a liquefaction-resistant total bacterial count test strip. The test strip comprises a transparent upper membrane and a base plate stacked on top of each other. The transparent upper membrane is provided with a liquefaction-resistant cold hydrogel and an anti-liquefaction additive. The base plate is provided with a mixture of a total bacterial count colorimetric culture medium and a compounded cold hydrogel powder.

[0024] The liquefaction-resistant cold hydrogel is a high-acyl gelling gel; the anti-liquefaction additive is glycine, and the dosage is 2% to 10% of the cold hydrogel powder.

[0025] The total colony count chromogenic medium consists of the following components by weight percentage: 40%–60% tryptone, 10%–20% yeast extract, 5%–10% glucose, 10%–20% sodium chloride, 2%–5% sodium pyruvate, and 0.1%–0.4% TTC.

[0026] The compound cold water gel is composed of guar gum and sodium carboxymethyl cellulose, with a ratio of 5:(5-8) between the two cold water soluble gel matrices.

[0027] The total colony count colorimetric culture medium and the compound cold hydrogel are mixed in a weight ratio of (1-2):(1-4).

[0028] This invention also proposes a method for preparing a total bacterial count test strip for liquefaction prevention, which includes the following steps:

[0029] (1) Preparation: liquefaction-resistant cold hydrogel, anti-liquefaction additive, total bacterial count colorimetric culture medium, and a mixed powder of the compound cold hydrogel; specifically including:

[0030] (1.1) Preparation of colorimetric medium for total bacterial count

[0031] Take 40%–60% tryptone, 10%–20% yeast extract, 5%–10% glucose, 10%–20% sodium chloride, 2%–5% sodium pyruvate and 0.1%–0.4% TTC respectively, grind them into powder, sieve them and mix the dry powders evenly for later use.

[0032] (1.2) Preparation of composite cold hydrogel powder

[0033] Take guar gum and sodium carboxymethyl cellulose, grind them into powder, sieve them, weigh them in a ratio of 5: (5-8) and mix them thoroughly.

[0034] (1.3) Preparation of mixed powder

[0035] Mix the total colony count colorimetric medium from step (1.1) with the compound cold hydrogel from step (1.2) thoroughly at a weight ratio of (1-2):(1-4).

[0036] (1.4) Preparation of anti-liquefaction cold hydrogel powder

[0037] Grind the high-acyl gel gel into powder and sieve it; grind glycine into powder and sieve it; the amount of glycine added is 2% to 10% of the weight of the gel gel powder; mix the two powders in the above proportion evenly and set aside.

[0038] (2) Spray the anti-liquefaction cold hydrogel powder at a spraying rate of 0.15-0.30 g / sheet on the inside of the transparent upper film (i.e., the side of the PET upper film facing the PET plastic base); spray the mixed powder containing the total bacterial count colorimetric culture medium and the compound cold hydrogel at a spraying rate of 0.05-0.20 g / sheet on the inside of the base plate (i.e., the side of the PET plastic base plate facing the PET upper film);

[0039] The transparent upper film is laminated and bonded to the base plate;

[0040] (3) Sterilization.

[0041] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention.

[0042] Example 1: Screening of anti-liquefaction cold water gels

[0043] Extensive experiments were conducted to initially screen common cold hydrogels, selecting those with high gel strength, fast gelation speed, and good water absorption. After initial screening, the selected monomeric gels were compounded in different proportions to identify compound gels with good gelation effects. The cold hydrogels from the initial and secondary screenings were compared to select monomeric gels or compound cold hydrogels that exhibited liquefaction resistance, fast gelation speed, and high gel strength. Ultimately, high-acyl gellan gum and xanthan gum were determined to have the best liquefaction resistance, followed by a compound gel of guar gum:carboxymethyl cellulose = 5:(5-8). However, xanthan gum inhibits bacterial growth; therefore, high-acyl gellan gum was selected as the cold hydrogel.

[0044] Example 2: Selection of Anti-liquefaction Additives

[0045] After extensive experimentation, metal cations (K) were selected. + Mg 2+ Ca 2+ The anti-liquefaction effects of Bacillus subtilis bacterial suspension were screened using gelling agents (potassium tetraborate, feed binder), and amino acids (threonine, tryptophan, glycine, lysine). The results are shown in Table 1. Mg was ultimately selected as the preferred anti-liquefaction agent. 2+ Threonine and glycine can enhance the coagulation power and decomposition resistance of cold hydrogels, and reduce liquefaction. Since glycine can also promote the rapid growth of bacteria, glycine was selected as an anti-liquefaction additive.

[0046] Table 1. Test results of different anti-liquefaction additives

[0047]

[0048] Example 3: Preparation of Anti-liquefaction Total Bacterial Count Test Strip

[0049] (1) Preparation of substrate adhesion mixture powder: Grind 20g tryptone, 5g yeast powder, 5g sodium chloride, 2.5g glucose, 1g sodium pyruvate, and 0.2g TTC into powder, pass through a 120-160 mesh sieve, and mix the dry powder thoroughly. This is the total colony count colorimetric culture medium powder. Grind 15g guar gum and 15g sodium carboxymethyl cellulose into powder, pass through a 120-160 mesh sieve, and mix thoroughly. This is the compound cold hydrogel powder. Take 30g of the above colorimetric culture medium powder and 30g of the compound cold hydrogel powder, and mix them thoroughly.

[0050] (2) Preparation of anti-liquefaction cold water gel: Take 60g of high acyl gel gel that has passed through a 120-mesh sieve, add 3.6g of glycine that has passed through a 120-mesh sieve, and mix evenly;

[0051] (3) Preparation of total bacterial count test strips: The above-mentioned anti-liquefaction hydrogel powder is evenly sprayed onto the inside of a tacky transparent PET upper film (thickness 0.05mm, length * width = 100 * 70cm), with a spraying amount of 0.20g / sheet; the above-mentioned mixed powder is evenly sprayed onto the inside of a tacky white PET plastic base plate (thickness 0.10mm, length * width = 100 * 70cm), with a spraying amount of 0.15g / sheet. The upper transparent film and the plastic base plate are sealed and bonded together with double-sided tape or pressure-sensitive adhesive, with a bonding width of 10mm.

[0052] (4) After production, package and sterilize with ethylene oxide.

[0053] Example 4: Preparation of Anti-liquefaction Total Bacterial Count Test Strip

[0054] (1) Preparation of substrate adhesion mixture powder: Grind 20g tryptone, 5g yeast powder, 5g sodium chloride, 2.5g glucose, 1g sodium pyruvate, and 0.2g TTC into powder, pass through a 120-160 mesh sieve, and mix the dry powder thoroughly. This is the total colony count colorimetric culture medium powder. Grind 15g guar gum and 24g sodium carboxymethyl cellulose into powder, pass through a 120-160 mesh sieve, and mix thoroughly. This is the compound cold hydrogel powder. Take 5g of the above colorimetric culture medium powder and 20g of the compound cold hydrogel powder, and mix them thoroughly.

[0055] (2) Preparation of anti-liquefaction cold hydrogel: Take 60g of high acyl gellan gum that has passed through a 120-mesh sieve, add 1.2g of glycine that has passed through a 120-mesh sieve, and mix evenly;

[0056] (3) Preparation of total bacterial count test strips: The above-mentioned anti-liquefaction hydrogel powder is evenly sprayed onto the inside of a tacky transparent PET upper film (thickness 0.05mm, length * width = 100 * 70cm), with a spraying amount of 0.20g / sheet; the above-mentioned mixed powder is evenly sprayed onto the inside of a tacky white PET plastic base plate (thickness 0.10mm, length * width = 100 * 70cm), with a spraying amount of 0.15g / sheet. The upper transparent film and the plastic base plate are sealed and bonded together with double-sided tape or pressure-sensitive adhesive, with a bonding width of 10mm.

[0057] (4) After production, package and sterilize with ethylene oxide.

[0058] Example 5: Preparation of Anti-liquefaction Total Bacterial Count Test Strip

[0059] (1) Preparation of substrate adhesion mixture powder: Grind 20g tryptone, 5g yeast powder, 5g sodium chloride, 2.5g glucose, 1g sodium pyruvate, and 0.2g TTC into powder, pass through a 120-160 mesh sieve, and mix the dry powder thoroughly. This is the total colony count colorimetric culture medium powder. Grind 15g guar gum and 18g sodium carboxymethyl cellulose into powder, pass through a 120-160 mesh sieve, and mix thoroughly. This is the compound cold hydrogel powder. Take 30g of the above colorimetric culture medium powder and 15g of the compound cold hydrogel powder, and mix them thoroughly.

[0060] (2) Preparation of anti-liquefaction cold hydrogel: Take 60g of high acyl gellan gum that has passed through a 120-mesh sieve, add 6.0g of glycine that has passed through a 120-mesh sieve, and mix evenly;

[0061] (3) Preparation of total bacterial count test strips: The above-mentioned anti-liquefaction hydrogel powder is evenly sprayed onto the inside of a tacky transparent PET upper film (thickness 0.05mm, length * width = 100 * 70cm), with a spraying amount of 0.20g / sheet; the above-mentioned mixed powder is evenly sprayed onto the inside of a tacky white PET plastic base plate (thickness 0.10mm, length * width = 100 * 70cm), with a spraying amount of 0.15g / sheet. The upper transparent film and the plastic base plate are sealed and bonded together with double-sided tape or pressure-sensitive adhesive, with a bonding width of 10mm.

[0062] (4) After production, package and sterilize with ethylene oxide.

[0063] Example 6: Application of the Anti-liquefaction Total Bacterial Count Test Strip

[0064] Referring to GB 4789.2-2022, take 25g (mL) of sample and place it in a sterile homogenizing cup or bag containing 225mL of phosphate buffer (or physiological saline). Homogenize for 1-2 minutes to prepare a 1:10 sample homogenate. If necessary, adjust the pH of the sample homogenate to 6.8-7.2 with sterile 1mol / L NaOH or 1mol / L HCl solution. Use a 1mL sterile pipette or pipette to draw 1mL of the 1:10 homogenate and inject it into a test tube containing 9mL of diluent. Shake to obtain a 1:100 sample homogenate. Repeat this process to prepare 10-fold serial dilutions of the sample homogenate. Use a 1mL sterile pipette or pipette tip for each additional dilution.

[0065] Based on the estimated contamination level of the samples, select 1–3 suitable dilutions of sample homogenate (liquid samples may include the undiluted solution) for testing. Place the test strip on a sterile, flat work surface, peel off the top membrane, and use a sterile pipette to add 1 mL of sample homogenate to the center of the test strip. Slowly cover the top membrane, taking care to avoid generating air bubbles during this process. Let it stand for about 2 minutes to allow the culture medium to solidify. Inoculate two strips for each dilution. Simultaneously, add 1 mL of blank dilution to two sterile test strips as blank controls.

[0066] After inoculation, place the test strip horizontally upwards in a constant temperature incubator with the transparent side facing up. Incubate at 36±1℃ for 24–48 hours, and for aquatic products at 30±1℃ for 72±3 hours. Results can then be read. Count all red colonies, regardless of size or clarity. The appropriate counting range is 30 CFU–300 CFU.

[0067] Example 7: Evaluation of the effect of anti-liquefaction bacterial count test strip.

[0068] Comparison of test results of standard bacterial suspensions

[0069] Standard control bacteria for total bacterial count, coliform bacteria, Escherichia coli, Staphylococcus aureus, Salmonella, Bacillus subtilis, Bacillus licheniformis magnetic beads (isolated, purified, and preserved in our laboratory), Bacillus polymyxa magnetic beads (isolated, purified, and preserved in our laboratory), Bacillus coagulans magnetic beads (isolated, purified, and preserved in our laboratory), Bacillus megaterium magnetic beads (isolated, purified, and preserved in our laboratory), and Bacillus subtilis magnetic beads (isolated, purified, and preserved in our laboratory) were cultured in nutrient broth for 16–18 h. The bacterial suspension was then diluted with sterile phosphate diluent to a suitable concentration, approximately 50–150 CFU / mL. 1 mL of the sample solution was added to the center of the test slide, and a thin film was slowly placed on top. The slide was allowed to stand for 2 min before incubation. Simultaneously, 3M (Petrifilm 6406, Comparative Example 1) and MicroFast LR1001 (Comparative Example 2) were used as controls. The samples were incubated at 36±1℃ for 24 h, and the results were observed. Figure 1-3 As shown in Figure 1 and Table 2, the results indicate that the test strip of this invention has a significantly better anti-spore-forming effect than the total bacterial count test strips of Petrifilm 6406 and MicroFastLR1001.

[0070] Table 2 Comparison of Standard Bacterial Suspension Detection Results

[0071]

[0072] 2) Actual sample testing results

[0073] Different food samples were prepared into appropriate dilutions according to "Example 6". 1 ml of each sample solution was used for total bacterial count using test strips, and PCA plate counting was performed simultaneously according to GB 4789.2-2022. The experiment was repeated twice, and the average results were taken (see Table 3). The linear correlation curves of the two methods are shown in [Table 3]. Figure 4 As shown in the table and graph, the coefficient of variation (CV) between the test strip method and the national standard method is <5%, and the agreement rate is >96.97%. The total bacterial count test strip and the national standard method show a good linear relationship: y = 0.033 + 0.974x, R0. 2 =0.997, indicating that there is no significant difference between the two detection methods in the counting results of all samples, including standard strains and actual samples.

[0074] Table 3 shows the total bacterial count of different solutions detected using the test strip method.

[0075]

[0076]

[0077] Unless otherwise specified, the raw materials and equipment used in this invention are all commonly used in the field; unless otherwise specified, the methods used in this invention are all conventional methods in the field.

[0078] Unless otherwise specified, the terms used in this specification have the same meaning as those commonly understood by those skilled in the art; however, in the event of any conflict, the definitions in this specification shall prevail.

[0079] The terms “comprising,” “including,” “containing,” “having,” “comprising,” or other variations thereof are intended to cover non-closed inclusion, and no distinction is made between these terms. The term “comprising” means that other steps and ingredients may be added without affecting the final result. The term “comprising” also includes the terms “consisting of” and “substantially consisting of”. The compositions and methods / processes of the present invention comprise, consist of, and substantially consist of the essential elements and limitations described herein, as well as any additional or optional ingredients, components, steps, or limitations described herein.

[0080] All numerical values ​​or expressions relating to component amounts, process conditions, etc., used in the specification and claims are to be understood to be modified with “about” in all cases. All ranges relating to the same component or property include endpoints that can be independently combined. Because these ranges are continuous, they include every value between the minimum and maximum values. It should also be understood that any numerical range referenced in this application is intended to include all subranges within that range.

[0081] The above embodiments merely illustrate the implementation of the present invention, and their descriptions are relatively specific and detailed. However, they should not be construed as limiting the scope of the present invention. Any technical solutions obtained by adopting equivalent substitutions or equivalent transformations should fall within the protection scope of the present invention.

Claims

1. A liquefaction-resistant total bacterial count test strip, the test strip comprising a transparent upper film and a base plate stacked on top of each other, characterized in that, The transparent upper membrane is provided with liquefaction-resistant cold hydrogel and anti-liquefaction additives, and the bottom plate is provided with a mixture of colony count colorimetric culture medium and compound cold hydrogel powder. The liquefaction-resistant cold hydrogel is a high-acyl gelling gel; the anti-liquefaction additive is glycine, and the dosage is 2% to 10% of the cold hydrogel powder. The compound cold hydrogel is composed of guar gum and sodium carboxymethyl cellulose by weight; the weight ratio of guar gum to sodium carboxymethyl cellulose is 5:(5~8). The total colony count colorimetric culture medium and the compound cold hydrogel are mixed in a weight ratio of (1~2):(1~4).

2. The anti-liquefaction total bacterial count test strip according to claim 1, characterized in that, The total colony count chromogenic medium is composed of the following components by weight percentage: 40%–60% tryptone, 10%–20% yeast extract, 5%–10% glucose, 10%–20% sodium chloride, 2%–5% sodium pyruvate, and 0.1%–0.4% TTC.

3. A method for preparing an anti-liquefaction bacterial count test strip as described in claim 1, characterized in that, It includes the following steps: (1) Preparation: liquefaction-resistant cold hydrogel, anti-liquefaction additive, total bacterial count colorimetric culture medium and mixed powder of compound cold hydrogel; (2) Spray the liquefaction-resistant cold hydrogel and anti-liquefaction additive on the inside of the transparent upper membrane; spray the mixed powder of the total colony count colorimetric culture medium and the compound cold hydrogel on the inside of the base plate; The transparent upper film is laminated and bonded to the base plate; (3) Sterilization.

4. The method for preparing the anti-liquefaction bacterial count test strip according to claim 3, characterized in that, The spraying amount of the liquefaction-resistant cold hydrogel and the anti-liquefaction additive is 0.15~0.30 g / sheet; the spraying amount of the total colony count colorimetric culture medium and the mixed powder of the compound cold hydrogel is 0.05~0.20 g / sheet.

5. The method for preparing the anti-liquefaction bacterial count test strip according to claim 3, characterized in that, Step (1) specifically includes: (1.1) Preparation of colorimetric culture medium for total bacterial count Take 40%~60% tryptone, 10%~20% yeast extract, 5%~10% glucose, 10%~20% sodium chloride, 2%~5% sodium pyruvate and 0.1%~0.4% TTC respectively, grind them into powder, sieve them and mix the dry powders evenly for later use; (1.2) Preparation of compound cold hydrogel powder Take guar gum and sodium carboxymethyl cellulose separately, grind them into powder, sieve them, weigh them in a ratio of 5: (5~8) and mix them thoroughly. (1.3) Preparation of mixed powder Mix the total colony count colorimetric medium from step (1.1) with the compound cold hydrogel from step (1.2) thoroughly at a weight ratio of (1~2):(1~4); (1.4) Preparation of anti-liquefaction cold hydrogel powder Grind the high-acyl gellan gel into powder and sieve it; grind glycine into powder and sieve it; the amount of glycine added is 2% to 10% of the weight of the gellan gel powder; mix the two powders in the above proportion evenly and set aside.

6. The application of the anti-liquefaction total bacterial count test strip as described in claim 1 in detecting the total bacterial count in food.

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