Sample processing method for cervical exfoliated cell pcr detection

Abstract

The application relates to a sample processing method for cervical exfoliative cell PCR detection, which adopts a sample processing reagent, including a processing liquid, a protection liquid, a conversion liquid, a first purification liquid, a second purification liquid, a third purification liquid and a collection liquid. The processing liquid contains 0.1-0.5N sodium hydroxide and 4-8mol / L urea; the protection liquid contains 1-8mM guanidine isothiocyanate, 0.1-5% SDS, 5-150mM Tris-Hcl and 5-100mM EDTA; and the conversion liquid contains 100-500mM sodium bisulfite, 50-250mM ammonium sulfite, 2-15M ammonium bisulfite, 0.1-3M tetrahydrofurfuryl alcohol, 1-5mM TEPA and 0.1-2M magnesium sulfite. The sample is lysed and protected by the processing liquid and the protection liquid, direct conversion and detection are facilitated, processing time and steps are saved, DNA loss is reduced, cost is saved, the accuracy and reliability of detection results are ensured.

Description

Technical Field

[0001] This invention relates to a sample processing method for PCR detection of cervical exfoliated cells, applicable to the field of in vitro biomedical detection technology. Background Technology

[0002] Cervical cancer is a common malignant tumor of the female reproductive system that occurs in the cervix, posing a significant threat to women's health and lives. Early diagnosis significantly improves the detection rate and patient survival rate in the prevention and treatment of cervical cancer. Currently, early screening for cervical cancer commonly uses liquid-based cytology, colposcopy, and histopathology combined with HPV testing. However, these methods have several limitations. For example, liquid-based cytology is heavily influenced by subjective factors, cannot differentiate the grade of cervical cell lesions, and the overlap of blood and mucus cells can affect the identification of abnormal cells, resulting in a high false-negative rate. Histopathology and colposcopy are not only difficult to sample but can also damage the cervix. HPV testing has high sensitivity but low specificity; a positive result only indicates a transient infection, and its value in predicting the occurrence and progression of cervical cancer needs further confirmation.

[0003] With the continuous development of biotechnology, and given the shortcomings of existing cervical cancer detection methods, epigenetics and epigenomics, represented by methylation analysis, are increasingly recognized for their role in tumor detection. Methods such as PCR (Polymerase Chain Reaction), gene sequencing, and circulating cell-free DNA detection have shown good efficacy in the early diagnosis of cervical cancer. However, most of these methods are based on genotyping after nucleic acid extraction and amplification. Traditional nucleic acid extraction requires purification, washing, drying, and elution after sample lysis before transformation and detection. The entire process is complex, prone to errors and contamination, and takes approximately 8 hours from sample extraction to instrumental testing, which is time-consuming.

[0004] Existing technologies also offer numerous time-saving methods, such as the approach employed in the published patent application number 201910405182.3. This method simultaneously performs sample lysis and magnetic bead adsorption of nucleic acids, using magnetic beads as a carrier to adsorb the lysed DNA, achieving preliminary purification and separation. This replaces the purification, rinsing, drying, and elution processes in traditional methods, thereby shortening extraction and transformation time. However, using magnetic bead adsorption instead of purification has significant limitations: First, a large amount of nucleic acid DNA is lost during extraction and transformation on carriers such as magnetic beads, affecting not only the reliability of subsequent PCR results but also increasing detection costs. Second, directly placing the magnetic beads adsorbed with nucleic acid DNA under high-temperature (80~100℃) conditions for transformation can negatively impact the adsorption effect of the magnetic beads, thus affecting the DNA transformation effect and reducing the accuracy of subsequent PCR results.

[0005] For example, patent application number 202210302663.3 discloses a nucleic acid extraction reagent and method for dried blood spot samples on filter paper. This patent describes a method of treating dried blood spot samples with alkaline lysis to obtain a sample suitable for PCR detection. However, due to the differences between cervical exfoliated cell samples and dried blood spot samples on filter paper, the disclosed method can easily damage the cell samples when used to process cervical exfoliated cells, resulting in insufficient extraction of nucleic acid DNA. This can lead to errors in subsequent PCR detection results and misjudgments. Summary of the Invention

[0006] To address the shortcomings of existing technologies, this invention proposes a sample processing method for PCR detection of cervical exfoliated cells. The technical solution adopted in this invention is as follows: a sample processing method for PCR detection of cervical exfoliated cells, employing sample processing reagents for PCR detection of cervical exfoliated cells. The processing reagents include a processing solution, a protective solution, a transformation solution, a first purification solution, a second purification solution, a third purification solution, and a collection solution. The processing solution includes 0.1-0.5N sodium hydroxide and 4-8 mol / L urea. The protective solution includes 1-8 mM guanidine isothiocyanate, 0.1-5% SDS, 5-150 mM Tris-HCl, and 5-100 mM EDTA. The transformation solution includes 100-500 mM sodium bisulfite, 50-250 mM ammonium sulfite, 2-15 M ammonium bisulfite, 0.1-3 M tetrahydrofurfuryl alcohol, 1-5 mM TEPA, and 0.1-2 M magnesium sulfite.

[0007] The processing methods include:

[0008] S1. Add the processing solution to the sample, vortex mix, then add the protective solution, vortex mix again to obtain the first product.

[0009] S2. Add conversion buffer to the first product, vortex to mix, and then place in a PCR instrument for conversion to obtain the second product.

[0010] S3. Add the first purification solution containing magnetic beads to the second product, mix well, and let stand at room temperature for 1 to 6 minutes. Then place it on a magnetic rack for magnetic separation for 1 to 10 minutes and discard the supernatant.

[0011] S4. Add the second purification solution to the product obtained in step S3 for rinsing. The rinsing method is to vortex and then place the product on a magnetic rack for 1 to 6 minutes, and then remove the supernatant.

[0012] S5. Add the third purification solution to the product obtained in step S4, vortex and incubate at room temperature, then place it on a magnetic rack for 1-6 minutes, and then remove the supernatant.

[0013] S6. Add the second purification solution to the product obtained in step S5, and rinse according to the rinsing method in step S4. Then repeat the current step once to obtain the third product.

[0014] S7. Discard all remaining liquid in the third product and air dry at room temperature. Then add the collected liquid, let it stand at room temperature, and place it on a magnetic rack for magnetic separation for 1-6 minutes. Aspirate the supernatant to obtain the required sample.

[0015] Furthermore, the first purification solution includes 1-6 mM guanidine isothiocyanate, 0.1-0.4% ethylene glycol ethyl ether acetate, 0.6-1.5 M guanidine hydrochloride, and sterile water. The magnetic beads are hydroxyl magnetic beads, and the content of the magnetic beads in the first purification solution is 5-15 mg / mL.

[0016] Furthermore, the second purification solution includes 70-90% ethanol and sterile water.

[0017] Furthermore, the third purification solution includes 100-300 mM sodium hydroxide, 50-150 mM sodium chloride, 1-10% glycerol, and 10-60% ethanol.

[0018] Furthermore, the collected solution includes 1~20mM Tris, 0.05~0.15mM EDTA, and sterile water.

[0019] Further, in step S1, the ratio of the sample volume to the processing liquid volume is 0.1~1, and the ratio of the protective liquid volume to the processing liquid volume is 0.1~2.

[0020] Furthermore, in step S2, the ratio of the volume of the first product to the volume of the conversion liquid is 6.5 to 20.

[0021] Furthermore, in step S2, the transformation conditions for transformation in the PCR instrument are as follows: first react at 95~101℃ for 2~12 minutes, then react at 54~62℃ for 30~80 minutes, and finally store at 4℃.

[0022] Furthermore, the volume ratio of the second product to the volume of the first purification solution is 0.2 to 0.3, the volume ratio of the second product to the volume of the second purification solution is 0.25 to 0.375, and the volume ratio of the second product to the volume of the third purification solution is 0.75 to 1.5.

[0023] Furthermore, in step S5, the incubation time at room temperature is 10-20 minutes.

[0024] Due to the application of the above technical solution, the present invention has the following advantages compared with the prior art:

[0025] This invention provides a sample processing method for PCR detection of cervical exfoliated cells. After lysing and protecting the cervical exfoliated cells with a treatment solution and a protective solution, the cells can be directly transformed using a transformation solution to obtain the sample required for PCR methylation detection. This eliminates the purification, rinsing, drying, and elution processes after lysis in traditional methods, significantly reducing the overall processing time, simplifying the steps, and lowering the possibility of errors and contamination. Furthermore, the extraction process eliminates the need for DNA adsorption via magnetic beads or other carriers, reducing DNA loss, saving costs, and ensuring the reliability of PCR results. It also avoids the adverse effects of high temperatures on the carrier during transformation, ensuring transformation efficiency and the accuracy of PCR results. Moreover, the treatment of cervical exfoliated cells with the treatment and protective solutions ensures sample integrity, further improving the accuracy of subsequent PCR results and preventing misinterpretations. Attached Figure Description

[0026] The following sections will describe some specific embodiments of the invention in a detailed manner, by way of example and not limitation, with reference to the accompanying drawings. The same reference numerals in the drawings denote the same or similar components or parts. Those skilled in the art should understand that these drawings are not necessarily drawn to scale. In the drawings:

[0027] Figure 1 This is the PAX1 amplification curve of Example 1 in this invention;

[0028] Figure 2 This is the amplification curve of EPB41L3 in Example 1 of this invention;

[0029] Figure 3 This is the amplification curve of FAM19A4 in Example 1 of this invention;

[0030] Figure 4 This is the ZNF582 amplification curve of Example 1 in this invention;

[0031] Figure 5 This is a graph showing the methylation detection results of Example 1 in this invention;

[0032] Figure 6 This is a graph showing the methylation detection results of Example 2 in this invention;

[0033] Figure 7 This is a graph showing the methylation detection results of Example 3 in this invention;

[0034] Figure 8 This is a comparison chart of the CT values ​​of the internal reference gene ACTB1 in Example 1 and the control example of this invention;

[0035] Figure 9 This is a comparison chart of the CT values ​​of the PAX1 gene in Example 1 and the control example of this invention;

[0036] Figure 10 This is a comparison chart of the CT values ​​of the EPB41L3 gene in Example 1 and the control example of this invention;

[0037] Figure 11 This is a comparison chart of the CT values ​​of the internal reference gene ACTB2 in Example 1 and the control example of this invention;

[0038] Figure 12 This is a comparison chart of the CT values ​​of the FAM19A4 gene in Example 1 and the control example of this invention;

[0039] Figure 13 This is a comparison chart of the CT values ​​of the ZNF582 gene in Example 1 and the control example of this invention. Implementation

[0040] The technical solution of the present invention will now be clearly and completely described with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0041] Furthermore, the technical features involved in the different embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other. Example

[0042] See attached document Figures 1-5This embodiment provides a sample processing method for PCR detection of cervical exfoliated cells, using sample processing reagents for PCR detection of cervical exfoliated cells. The processing reagents include a processing solution, a protection solution, a transformation solution, a first purification solution, a second purification solution, a third purification solution, and a collection solution. The treatment solution includes 0.2N sodium hydroxide and 6mol / L urea; the protection solution includes 4mM guanidine isothiocyanate, 0.5% SDS, 80mM Tris-HCl, and 55mM EDTA; the conversion solution includes 320mM sodium bisulfite, 140mM ammonium sulfite, 9M ammonium bisulfite, 1M tetrahydrofurfuryl alcohol, 2mM TEPA, and 0.6M magnesium sulfite; the first purification solution includes 4mM guanidine isothiocyanate, 0.3% ethylene glycol ethyl ether acetate, 1M guanidine hydrochloride, and sterile water; the second purification solution includes 80% ethanol and sterile water; the third purification solution includes 200mM sodium hydroxide, 100mM sodium chloride, 5% glycerol, and 40% ethanol; and the collection solution includes 10mM Tris, 0.1mM EDTA, and sterile water.

[0043] Seven cervical exfoliated cell samples were selected for testing. The sample numbers were: LDT127, LDT130, LDT152, LDT154, LDT158, LDT160, and LDT163. Among these, three samples (LDT127, LDT130, and LDT160) showed CIN III on colposcopy, while four samples (LDT152, LDT154, LDT158, and LDT163) showed chronic inflammation on colposcopy. All seven samples were processed using the method of this invention. The processing method included:

[0044] S1. Place the exfoliated cervical cells in the cell preservation solution and vortex at the highest speed to obtain a sample. Take 2 ml of the sample into a centrifuge tube, centrifuge at 12000 rpm for 2 minutes, discard the supernatant, then add 40 μL of processing solution to the sample, vortex for 5 minutes, add 40 μL of preservation solution, vortex again to obtain the first product.

[0045] S2. Take 40 μL of the first product into a PCR tube, add 110 μL of conversion buffer, vortex to mix, and then place in a PCR instrument for conversion to obtain the second product. Specifically, the conversion conditions are: react at 98℃ for 8 minutes, react at 54℃ for 60 minutes, and then store at 4℃.

[0046] S3. Add 600µL of the first purification solution containing magnetic beads to the second product, mix well, let stand at room temperature for 2 minutes, then place on a magnetic rack for magnetic separation for 5 minutes, and discard the supernatant; wherein, the magnetic beads are hydroxyl magnetic beads, and the content of magnetic beads in the first purification solution is 10mg / mL.

[0047] S4. Add 200µL of the second purification solution to the product obtained in step S3 for rinsing. The rinsing method is to vortex and then place the product on a magnetic rack for 2 minutes, and then remove the supernatant.

[0048] S5. Add 150µL of the third purification solution to the product obtained in step S4, vortex and incubate at room temperature for 12 minutes, then place it on a magnetic rack for 2 minutes, and then remove the supernatant.

[0049] S6. Add 200µL of the second purification solution to the product obtained in step S5, and rinse according to the rinsing method in step S4. Then repeat the current step once to obtain the third product.

[0050] S7. Discard all remaining liquid in the third product and air dry at room temperature. Then add 22µL of the collection solution, let stand at room temperature for 3 minutes, and then place it on a magnetic rack for magnetic separation for 3 minutes. Aspirate the supernatant to obtain the required sample.

[0051] The methylation levels of seven samples treated with the above method were detected using PCR reaction solutions of cervical cancer markers PAX1, ZNF582, EPB41L3, and FAM19A4 genes, and the results were interpreted based on the ΔCt between each gene and the ACTB gene.

[0052] The primer pairs for detecting PAX1 gene methylation are shown in SEQ ID No. 1 and SEQ ID No. 2; the probe sequence for PAX1 gene methylation is shown in SEQ ID No. 3; the primer pairs for detecting EPB41L3 gene methylation are shown in SEQ ID No. 4 and SEQ ID No. 5; the probe sequence for detecting EPB41L3 gene methylation is shown in SEQ ID No. 6; the primer pairs for detecting FAM19A4 gene methylation are shown in SEQ ID No. 7 and SEQ ID No. 8; the probe sequence for detecting FAM19A4 gene methylation is shown in SEQ ID No. 9; the primer pairs for detecting ZNF582 gene methylation are shown in SEQ ID No. 10 and SEQ ID No. 11; the probe sequence for detecting ZNF582 gene methylation is shown in SEQ ID No. 12; and the primer pairs for detecting ACTB gene methylation are shown in SEQ ID No. 13 and SEQ ID No. 14. The sequence shown in SEQ ID No. 14 is the probe sequence for detecting ACTB gene methylation, and the sequence shown in SEQ ID No. 15 is the probe sequence for this purpose. The specific primer and probe nucleotide sequences are shown in the table below:

[0053]

[0054] Methods for detecting methylation levels include:

[0055] ① Prepare PCR1 and PCR2 reaction solutions. The components and amounts of both are shown in the table below:

[0056]

[0057] ② Mix the PCR1 reaction solution thoroughly and incubate for 2 minutes. Then add it to an 8-tube PCR tube with a capacity of 200 μL. Aliquot 15 μL into each PCR tube and add 5 μL of the sample to be tested to obtain the first set of test samples. Aliquot the PCR2 reaction solution using the same method to obtain the second set of test samples.

[0058] ③ Select three channels (FAM, VIC, and CY5) to perform quantitative real-time PCR detection on the first and second groups of test samples, respectively. The passive reference was set to none. Specific reaction conditions are shown in the table below:

[0059]

[0060] ④ The difference ΔCt between the Ct value of any one of the PAX1, EPB41L3, FAM19A4, and ZNF582 genes and the Ct value of the ACTB gene is used to reflect the relative quantification between the detected gene and the ACTB internal reference gene. ΔCt is used as the criterion to determine the methylation level of the four genes (PAX1, EPB41L3, FAM19A4, and ZNF582). Specific results are attached. Figure 5 As shown. Example

[0061] See attached document Figure 6The difference between this embodiment and Example 1 is that the concentrations of the solutions in the treatment reagent are different, while the other treatment methods and methylation detection methods are the same as in Example 1. The processing reagents in this embodiment include: a processing solution comprising 0.1N sodium hydroxide and 4 mol / L urea; a protective solution comprising 1 mM guanidine isothiocyanate, 0.1% SDS, 5 mM Tris-HCl, and 5 mM EDTA; a conversion solution comprising 100 mM sodium bisulfite, 50 mM ammonium bisulfite, 2 M ammonium bisulfite, 0.1 M tetrahydrofurfuryl alcohol, 1 mM TEPA, and 0.1 M magnesium sulfite; a first purification solution comprising 1 mM guanidine isothiocyanate, 0.1% ethylene glycol ethyl ether acetate, 0.6 M guanidine hydrochloride, and sterile water; the magnetic bead content in the first purification solution is 5 mg / mL; a second purification solution comprising 70% ethanol and sterile water; a third purification solution comprising 100 mM sodium hydroxide, 50 mM sodium chloride, 1% glycerol, and 10% ethanol; and a collection solution comprising 1 mM Tris, 0.05 mM EDTA, and sterile water. The methylation detection results of the samples tested in this embodiment are attached. Figure 6 As shown. Example

[0062] See attached document Figure 7 The difference between this embodiment and Example 1 is that the concentrations of the solutions in the treatment reagent are different, while the other treatment methods and methylation detection methods are the same as in Example 1. The processing reagents in this embodiment include: a processing solution comprising 0.5N sodium hydroxide and 8 mol / L urea; a protective solution comprising 8 mM guanidine isothiocyanate, 5% SDS, 150 mM Tris-HCl, and 100 mM EDTA; a conversion solution comprising 500 mM sodium bisulfite, 250 mM ammonium bisulfite, 15 M ammonium bisulfite, 3 M tetrahydrofurfuryl alcohol, 5 mM MTEEPA, and 2 M magnesium sulfite; a first purification solution comprising 6 mM guanidine isothiocyanate, 0.4% ethylene glycol ethyl ether acetate, 1.5 M guanidine hydrochloride, and sterile water; the magnetic bead content in the first purification solution is 15 mg / mL; a second purification solution comprising 90% ethanol and sterile water; a third purification solution comprising 300 mM sodium hydroxide, 150 mM sodium chloride, 10% glycerol, and 60% ethanol; and a collection solution comprising 20 mM Tris, 0.15 mM EDTA, and sterile water. The methylation detection results of the samples tested in this embodiment are attached. Figure 7 As shown.

[0063] Comparison Example

[0064] See attached document Figures 8-13This control example used 7 cervical exfoliated cell samples, with sample numbers and colposcopy results identical to those in Example 1. Nucleic acid extraction was performed on the 7 cervical exfoliated cell samples using the extraction reagents and methods disclosed in patent application number 202210302663.3. The concentrations of each component in the extraction reagent were selected as the optimal concentrations disclosed in that patent. The resulting test samples were tested according to the methylation level detection method disclosed in Example 1, and the test results were compared with those of Example 1, as shown in the appendix. Figures 8-13 As shown.

[0065] See attached document Figures 5-7 Furthermore, based on the detection results of Examples 1-3, it can be seen that the methylation detection results of the 7 cervical exfoliated cell samples selected in Examples 1, 2, and 3, after being processed by the corresponding processing reagents and methods used in the examples, were 100% consistent with the colposcopy detection results. This proves the effectiveness of the sample processing method disclosed in this invention. In addition, Examples 1, 2, and 3 selected low, medium, and high concentration values ​​of each solution component in the processing reagent disclosed in this invention, and based on the detection results of Examples 1, 2, and 3, the effectiveness of the content range of each solution component in the processing reagent disclosed in this invention can be demonstrated.

[0066] Further, refer to the appendix Figures 8-13 It is evident that the CT values ​​of both the internal reference gene and the target gene in the samples processed using the method disclosed in this invention are lower than those in the control patent with application number 202210302663.3 during methylation detection. This demonstrates that the processing method of this invention not only achieves rapid and thorough DNA extraction but also ensures the integrity of the DNA strand, further improving transformation and recovery efficiency. Therefore, the effectiveness of the processing method disclosed in this invention is far superior to existing technologies, including the patent with application number 202210302663.3.

[0067] In summary, the sample processing method disclosed in this invention simplifies processing steps, reduces processing time, and ensures effectiveness.

[0068] Due to the application of the above technical solution, the present invention has the following advantages compared with the prior art:

[0069] This invention provides a sample processing method for PCR detection of cervical exfoliated cells. After lysing and protecting the cervical exfoliated cells with a treatment solution and a protective solution, the cells can be directly transformed using a transformation solution to obtain the sample required for PCR methylation detection. This eliminates the purification, rinsing, drying, and elution processes after lysis in traditional methods, significantly reducing the overall processing time, simplifying the steps, and lowering the possibility of errors and contamination. Furthermore, the extraction process eliminates the need for DNA adsorption via magnetic beads or other carriers, reducing DNA loss, saving costs, and ensuring the reliability of PCR results. It also avoids the adverse effects of high temperatures on the carrier during transformation, ensuring transformation efficiency and the accuracy of PCR results. Moreover, the treatment of cervical exfoliated cells with the treatment and protective solutions ensures sample integrity, further improving the accuracy of subsequent PCR results and preventing misinterpretations.

[0070] The above embodiments are only for illustrating the technical concept and features of the present invention. Their purpose is to enable those skilled in the art to understand the content of the present invention and implement it accordingly. They should not be used to limit the scope of protection of the present invention. All equivalent changes or modifications made in accordance with the spirit and essence of the present invention should be covered within the scope of protection of the present invention.

Claims

1. A sample processing method for PCR detection of cervical exfoliated cells, characterized in that, The sample processing reagents used for PCR detection of cervical exfoliated cells include a processing solution, a protective solution, a transformation solution, a first purification solution, a second purification solution, a third purification solution, and a collection solution. The processing solution includes 0.1-0.5N sodium hydroxide and 4-8 mol / L urea. The protective solution includes 1-8 mM guanidine isothiocyanate, 0.1-5% SDS, 5-150 mM Tris-HCl, and 5-100 mM EDTA. The transformation solution includes 100-500 mM sodium bisulfite, 50-250 mM ammonium bisulfite, 2-15 M ammonium bisulfite, 0.1-3 M tetrahydrofurfuryl alcohol, 1-5 mM MTEEPA, and 0.1-2 M magnesium sulfite. The processing method includes: S1. Add the processing liquid to the sample, vortex mix, then add the protective liquid, vortex mix again to obtain the first product. S2. Add the conversion solution to the first product, vortex to mix, and then place it in a PCR instrument for conversion to obtain the second product. The conversion conditions in the PCR instrument are as follows: first react at 95~101℃ for 2~12 minutes, then react at 54~62℃ for 30~80 minutes, and finally store at 4℃. S3. Add the first purified solution containing magnetic beads to the second product, mix well, let stand at room temperature for 1 to 6 minutes, then place on a magnetic rack for magnetic separation for 1 to 10 minutes, and discard the supernatant. S4. Add the second purification solution to the product obtained in step S3 for rinsing. The rinsing method is to vortex and then place the product on a magnetic rack for 1 to 10 minutes, and then remove the supernatant. S5. Add the third purification solution to the product obtained in step S4, vortex and incubate at room temperature, then place on a magnetic rack for 1-10 minutes, and then remove the supernatant. S6. Add the second purification solution to the product obtained in step S5, and rinse according to the rinsing method described in step S4. Then repeat the current step once to obtain the third product. S7. Discard all the remaining liquid in the third product and air dry it at room temperature. Then add the collected liquid, let it stand at room temperature, and place it on a magnetic rack for magnetic separation for 1 to 10 minutes. Aspirate the supernatant to obtain the required sample to be tested.

2. The sample processing method for PCR detection of cervical exfoliated cells according to claim 1, characterized in that, The first purification solution comprises 1-6 mM guanidine isothiocyanate, 0.1-0.4% ethylene glycol ethyl ether acetate, 0.6-1.5 M guanidine hydrochloride, and sterile water. The magnetic beads are hydroxyl magnetic beads, and the content of the magnetic beads in the first purification solution is 5-15 mg / mL.

3. The sample processing method for PCR detection of cervical exfoliated cells according to claim 1, characterized in that, The second purification solution includes 70-90% ethanol and sterile water.

4. The sample processing method for PCR detection of cervical exfoliated cells according to claim 1, characterized in that, The third purification solution comprises 100-300 mM sodium hydroxide, 50-150 mM sodium chloride, 1-10% glycerol, and 10-60% ethanol.

5. The sample processing method for PCR detection of cervical exfoliated cells according to claim 1, characterized in that, The collection solution includes 1~20mM Tris, 0.05~0.15mM EDTA, and sterile water.

6. The sample processing method for PCR detection of cervical exfoliated cells according to claim 1, characterized in that, In step S1, the ratio of the volume of the sample to the volume of the treatment liquid is 0.1 to 1, and the ratio of the volume of the protective liquid to the volume of the treatment liquid is 0.1 to 2.

7. The sample processing method for PCR detection of cervical exfoliated cells according to claim 1, characterized in that, In step S2, the ratio of the volume of the first product to the volume of the conversion liquid is 6.5 to 20.

8. The sample processing method for PCR detection of cervical exfoliated cells according to claim 1, characterized in that, The volume ratio of the second product to the volume of the first purified solution is 0.2 to 0.3, the volume ratio of the second product to the volume of the second purified solution is 0.25 to 0.375, and the volume ratio of the second product to the volume of the third purified solution is 0.75 to 1.

5.

9. The sample processing method for PCR detection of cervical exfoliated cells according to claim 1, characterized in that, In step S5, the incubation time at room temperature is 10-20 minutes.

Images