CN112798370A - Quality control material for detecting visible components of vaginal secretion and preparation method thereof - Google Patents
Quality control material for detecting visible components of vaginal secretion and preparation method thereof Download PDFInfo
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- 210000003743 erythrocyte Anatomy 0.000 claims abstract description 78
- 239000000126 substance Substances 0.000 claims abstract description 76
- 241000186660 Lactobacillus Species 0.000 claims abstract description 45
- 229940039696 lactobacillus Drugs 0.000 claims abstract description 45
- 241000222122 Candida albicans Species 0.000 claims abstract description 39
- 229940095731 candida albicans Drugs 0.000 claims abstract description 39
- 239000006285 cell suspension Substances 0.000 claims abstract description 33
- 210000000265 leukocyte Anatomy 0.000 claims abstract description 33
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- 238000000926 separation method Methods 0.000 claims abstract description 4
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- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 27
- 210000004027 cell Anatomy 0.000 claims description 23
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/34—Purifying; Cleaning
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/38—Diluting, dispersing or mixing samples
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/84—Systems specially adapted for particular applications
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N2001/2893—Preparing calibration standards
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Abstract
The invention discloses a quality control material for detecting visible components of vaginal secretion and a preparation method thereof, comprising a red blood cell quality control material, a white blood cell quality control material, an oral epithelial cell quality control material, a candida albicans quality control material and a lactobacillus quality control material, wherein the obtained red blood cells and white blood cells are respectively subjected to anticoagulation whole blood, and after separation and purification, the red blood cells are solidified and stored to obtain the red blood cell quality control material and the white blood cell quality control material; respectively subculturing the obtained oral epithelial cells, candida albicans and lactobacillus, and after centrifugal dilution, fixing and storing the diluted epithelial cell suspension to obtain the oral epithelial cell quality control substance, the candida albicans quality control substance and the lactobacillus quality control substance.
Description
Technical Field
The invention relates to the technical field of analysis of visible components of vaginal secretion, in particular to a quality control material for detecting the visible components of the vaginal secretion and a preparation method thereof.
Background
The examination of vaginal secretion is one of the routine items in clinical examinations of vagina, obstetrics and the like in modern medicine, and plays an important role in the diagnosis of diseases.
In order to reduce the load of the inspectors on the microscopic examination of the visible components of the vaginal secretion, improve the inspection process, reduce the errors caused by human factors and promote the standardization of the inspection of the visible components of the vaginal secretion, automatic visible component analyzers are continuously on the market. With the continuous application of the automatic instrument, how to make the instrument exert the maximum effect is very important, the rapid, accurate and timely inspection is provided for a clinical laboratory, the working efficiency is improved, and the quality control work of the instrument is improved. However, the quality control of visible components in the vaginal secretion is still lack of a relatively comprehensive quality control product, and basically only the quality control products of red blood cells and white blood cells are available. In addition, the prior art has complex separation and purification and solidification processes for the erythroblasts, and the conditions of cell aggregation, cell expansion, cell morphology shrinkage and the like exist, so that the product quality of the quality control material is greatly limited, and the accuracy of detecting the visible components of the vaginal secretion is reduced.
Disclosure of Invention
The invention aims to provide a quality control material for detecting visible components of vaginal secretion and a preparation method thereof, which improve the accuracy of detecting the visible components of the vaginal secretion.
In order to achieve the above object, in a first aspect, the present invention provides a method for preparing a quality control substance for detecting a visible component of a vaginal secretion, wherein the quality control substance for detecting a visible component of a vaginal secretion comprises a red blood cell quality control substance, a white blood cell quality control substance, an oral epithelial cell quality control substance, a candida albicans quality control substance and a lactobacillus quality control substance, comprising the following steps:
performing anticoagulation on the obtained red blood cells, separating and purifying, and then performing solidification storage on the red blood cells to obtain the red blood cell quality control substance;
separating and purifying the leucocytes after anticoagulation of the whole blood, and fixing and storing the qualified cell-coated suspension subjected to microscopic examination to obtain the leucocyte quality control substance;
subculturing the obtained oral epithelial cells, centrifugally diluting, and fixing and storing an epithelial cell suspension obtained by dilution to obtain the oral epithelial cell quality control substance;
culturing, centrifuging and microscopic examining the obtained candida albicans, and fixing and storing a diluted candida albicans suspension to obtain the candida albicans quality control substance;
and culturing, centrifuging and washing the obtained lactobacillus, and fixing and storing the diluted lactobacillus suspension to obtain the lactobacillus quality control substance.
Wherein, to the erythrocyte that obtains carry out anticoagulation whole blood, and after separating purification, will the erythrocyte solidifies the preservation, obtains erythrocyte quality control thing includes:
adding a hemolytic agent into the obtained erythrocyte sample to perform anticoagulation whole blood for 24 hours, centrifuging the anticoagulation whole blood, and diluting and washing the obtained erythrocyte layer for 3-6 times until microscopic examination is qualified to obtain erythrocyte suspension;
and solidifying and storing the erythrocyte suspension to obtain the erythrocyte quality control substance.
Wherein, the erythrocyte suspension is solidified and stored to obtain the erythrocyte quality control substance, which comprises:
fixing the fixing solution and the erythrocyte suspension in a volume ratio of 6:1 for 24 hours at the temperature of 36.5 ℃ and at the speed of 50 r/min, and washing the erythrocyte suspension without the fixing solution with physiological saline for three times to finish the first fixing;
and (3) obtaining the fixing solution again, fixing and washing again according to the method for the first fixing, adding a preservation solution into the erythrocyte suspension after the fixing is finished, and preserving at the temperature of 2-8 ℃.
Performing subculture on the obtained oral epithelial cells, performing centrifugal dilution, and fixing and storing an epithelial cell suspension obtained by dilution to obtain the oral epithelial cell quality control substance, wherein the method comprises the following steps:
obtaining oral epithelial cells, subculturing to the third generation, centrifuging and washing for 3-5 times until microscopic examination is qualified, and diluting with normal saline to obtain epithelial cell suspension;
and (3) fixing and storing the epithelial cell suspension to obtain the oral epithelial cell quality control substance.
Wherein, the epithelial cell suspension is fixed and preserved to obtain the oral epithelial cell quality control substance, which comprises:
fixing and washing the epithelial cell suspension and the fixing solution with the volume ratio of 6:1 for two times at 36.5 ℃ under the condition of 50 r/min;
and adding a preservation solution into the epithelial cell suspension after fixation is completed, and preserving at the temperature of 2-8 ℃.
Wherein, the fixing solution and the epithelial cell suspension with the volume ratio of 6:1 are fixed and washed twice under the conditions of 36.5 ℃ and 50 r/min, and the method comprises the following steps:
fixing the fixing solution and the erythrocyte suspension in a volume ratio of 6:1 for 24 hours at the temperature of 36.5 ℃ and at the speed of 50 r/min, and washing the epithelial cell suspension without the fixing solution by using physiological saline for three times to finish the first fixing;
re-obtaining the fixing solution, and fixing the fixing solution and the epithelial cell suspension after the first fixing for 6 hours at the temperature of 36.5 ℃ and at the speed of 50 r/min according to the volume ratio of 6: 1;
and (3) washing the epithelial cell suspension after the fixing solution is removed by using physiological saline three times to finish the second fixing of the epithelial cell suspension.
In a second aspect, the present invention provides a quality control material for detecting visible components of vaginal secretion, and the preparation method of the quality control material for detecting visible components of vaginal secretion according to the first aspect is applied to a quality control material for detecting visible components of vaginal secretion, and the quality control material for detecting visible components of vaginal secretion includes a red blood cell quality control material, a white blood cell quality control material, an oral epithelial cell quality control material, a candida albicans quality control material and a lactobacillus quality control material.
The quality control material for detecting the visible components of the vaginal secretion further comprises a hemolytic agent, a fixing solution and a preservation solution, wherein the hemolytic agent is a mixed solution of 1-2g/L sodium citrate, 0.3-2.7% triton-100 and 7% NaCl.
The quality control material for detecting the visible components of the vaginal secretion comprises a red blood cell quality control material, a white blood cell quality control material, an oral epithelial cell quality control material, a candida albicans quality control material and a lactobacillus quality control material, and after the obtained red blood cells are subjected to anticoagulation whole blood and are separated and purified, the red blood cells are solidified and stored to obtain the red blood cell quality control material; separating and purifying the leucocytes after anticoagulation of the whole blood, and fixing and storing the qualified cell-coated suspension subjected to microscopic examination to obtain the leucocyte quality control substance; subculturing the obtained oral epithelial cells, centrifugally diluting, and fixing and storing an epithelial cell suspension obtained by dilution to obtain the oral epithelial cell quality control substance; culturing, centrifuging and microscopic examining the obtained candida albicans, and fixing and storing a diluted candida albicans suspension to obtain the candida albicans quality control substance; the obtained lactobacillus is cultured, centrifuged and washed, and the diluted lactobacillus suspension is fixed and stored to obtain the lactobacillus quality control substance, except red blood cells and white blood cells, epithelial cells and the lactobacillus quality control substance are included, so that the variety of the quality control substance is enlarged, the product quality is greatly improved, and the accuracy of detecting visible components of vaginal secretion is improved.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, it is obvious that the drawings in the following description are only some embodiments of the present invention, and for those skilled in the art, other drawings can be obtained according to the drawings without creative efforts.
FIG. 1 is a schematic diagram of the steps of a method for preparing a quality control material for detecting visible components in vaginal discharge according to the present invention.
FIG. 2 is a picture under the microscope for wet quality control of red blood cells provided by the present invention.
FIG. 3 is a photograph under a microscope for wet quality control of fungi and epithelial cells provided by the present invention.
FIG. 4 is a photograph under a wet quality control microscope of leukocytes and epithelial cells provided by the present invention.
FIG. 5 is a picture of a microscope for wet quality control of fungal spores, blastospores and hyphae provided by the invention.
FIG. 6 is a photograph under a quality control microscope for dry staining of epithelial cells provided by the present invention.
FIG. 7 is a photograph under a quality control microscope for dry staining of leukocytes and erythrocytes provided by the present invention.
FIG. 8 is a photograph under a microscope for dry stain quality control of fungal spores and blastospores provided by the present invention.
FIG. 9 is a photograph under a dry staining quality control microscope of Lactobacillus provided by the present invention.
Detailed Description
Reference will now be made in detail to embodiments of the present invention, examples of which are illustrated in the accompanying drawings, wherein like or similar reference numerals refer to the same or similar elements or elements having the same or similar function throughout. The embodiments described below with reference to the drawings are illustrative and intended to be illustrative of the invention and are not to be construed as limiting the invention.
In the description of the present invention, "a plurality" means two or more unless specifically defined otherwise.
Referring to fig. 1, the present invention provides a method for preparing a quality control material for detecting visible components of vaginal secretion, wherein the quality control material for detecting visible components of vaginal secretion comprises a red blood cell quality control material, a white blood cell quality control material, an oral epithelial cell quality control material, a candida albicans quality control material and a lactobacillus quality control material, and the method comprises the following steps:
s101, performing anticoagulation on the obtained red blood cells, separating and purifying, and then performing solidification storage on the red blood cells to obtain the red blood cell quality control substance.
Specifically, the selection of the red blood cell sample: animal ox blood is selected, double-concave round cake-shaped cells without rupture, shrinkage and cell nucleus are selected as erythrocytes, and sodium citrate anticoagulation whole blood of ox is selected, preferably within 24 hours.
Separating and purifying red blood cells: anticoagulated whole blood is centrifuged, and after centrifugation, the whole blood is divided into three layers: sequentially arranging plasma, white blood cells and red blood cell layers from top to bottom, finally reserving the red blood cell layer, adding 4-7 times of physiological saline for dilution to form cell suspension, and continuously carrying out centrifugal washing to only reserve the red blood cells. Repeating for 3-6 times, performing microscopic examination to confirm that only pure red blood cells exist, and diluting with normal saline to obtain red blood cell suspension.
Fixing red blood cells: adding the erythrocyte suspension into 6 times of cell fixing solution, keeping the temperature constant at 36.5 ℃, shaking at low speed of 50 r/min for 24 hours, removing the fixing solution, washing with normal saline for 3 times to complete the first fixing, then, obtaining and adding the fixing solution again, keeping the temperature constant at 36.5 ℃, shaking at low speed of 50 r/min for 24 hours, washing with normal saline for 3 times, adding the cell storage solution, and storing at 2-8 ℃ for later use, wherein the final concentration is 50-1800 pieces/. mu.L.
S102, separating and purifying the leucocytes after anticoagulation of the whole blood, and fixing and storing the cell-coated suspension qualified by microscopic examination to obtain the leucocyte quality control substance.
Specifically, the hemolytic agent comprises the following components: 0.1mol/L phosphate buffer solution; the surfactant is phenoxyethanol, the pH value is 6.1-7.0, and the osmotic pressure is 230-.
The preparation process of the leucocyte quality control substance comprises the following steps:
selection of leukocyte samples: adding a hemolytic agent to the obtained leukocyte sample for 24 hours of anticoagulated whole blood, comprising:
the leucocyte is selected from animal bovine blood, cells with cell nucleus and spherical shape, and sodium citrate anticoagulated whole blood is selected, preferably for 24 hours;
separation and purification of leucocytes: centrifuging the anticoagulated whole blood, diluting the obtained leukocyte layer for 3-5 times by using physiological saline with the volume ratio of 6:1, adding hemolytic agent with three times of volume at 37 ℃, incubating for 1-2 times, centrifuging until the leukocyte is qualified by microscopic examination, and obtaining leukocyte suspension, wherein the anticoagulated whole blood specifically comprises the following components:
with the centrifugation of anticoagulation whole blood, the whole blood after the centrifugation is from last 3 layers down of dividing into: plasma, leucocyte and erythrocyte layer, reserving leucocyte layer, adding 6 times volume of physiological saline to form leucocyte suspension; centrifuging and washing for 3-5 times, diluting, adding 3 times of hemolytic agent, incubating at 37 deg.C for 10min, centrifuging, repeating for 1-2 times, removing excessive red blood cells, and performing microscopic examination to obtain leukocyte suspension without red blood cells.
And (3) fixing white blood cells: fixing and washing the fixing solution and the leukocyte suspension at the volume ratio of 6:1 for two times at the temperature of 36.5 ℃ and at the speed of 50 r/min; and adding a preserving fluid into the epithelial cell suspension after fixation, and preserving at the temperature of 2-8 ℃, specifically:
adding leukocyte suspension into 6 times volume of cell fixing solution, keeping the temperature at 36.5 deg.C, shaking at 50 rpm for 24 hr, removing fixing solution, washing with physiological saline for 3 times, adding fixing solution, keeping the temperature at 36.5 deg.C, shaking at 50 rpm for 6 hr, washing with physiological saline for 3 times, adding cell storage solution to the final concentration of 50-1800/μ L, and storing at 2-8 deg.C for use.
S103, subculturing the obtained oral epithelial cells, centrifugally diluting, and fixing and storing the diluted epithelial cell suspension to obtain the oral epithelial cell quality control substance.
Specifically, the oral epithelial cells were scraped off from the human mouth with a toothpick gently, subcultured, and left for use after subculture for 3 generations.
Centrifuging cultured oral epithelial cells, washing with normal saline, centrifuging, repeating for 3-5 times, and performing microscopic examination until epithelial cells are cleaned. Diluting with normal saline to obtain epithelial cell suspension.
Oral epithelial cell fixation: adding 6 times of cell fixing solution into the oral epithelial cell suspension, keeping the temperature at 36.5 ℃, shaking at low speed of 50 r/min for 24 hours, removing the fixing solution, washing with normal saline for 3 times, and completing the first fixing; adding the fixing solution, keeping the temperature constant at 36.5 ℃, shaking at low speed of 50 r/min for 6 hours, and washing with normal saline for 3 times to complete the second fixing; adding cell stock solution to the solution with final concentration of 50-500 pieces/. mu.L, and storing at 2-8 deg.C for use.
S104, culturing, centrifuging and microscopic examination are carried out on the obtained Candida albicans, and the diluted Candida albicans suspension is fixed and stored to obtain the Candida albicans quality control substance.
Specifically, the method comprises the steps of obtaining candida albicans, subculturing to the third generation, centrifuging and washing for 3-5 times until microscopic examination is qualified, and diluting by using normal saline to obtain a candida albicans suspension, wherein the candida albicans suspension comprises the following steps:
candida albicans is purchased, subcultured, and reserved for later use after subculture is carried out for 3 generations.
Centrifuging cultured Candida albicans for passage, washing with normal saline, centrifuging, repeating for 3-5 times, and performing microscopic examination until Candida albicans is washed. Diluting with normal saline to obtain Candida albicans suspension.
Fixing and washing the stationary liquid and the candida albicans suspension at the volume ratio of 6:1 for two times at the temperature of 36.5 ℃ and at the speed of 50 r/min; adding a preservation solution into the candida albicans suspension after fixation is completed, and preserving at the temperature of 2-8 ℃ to obtain the candida albicans quality control substance, which comprises the following steps:
and (3) candida albicans fixing: adding Candida albicans suspension into 6 times volume of cell fixing solution, keeping the temperature constant at 36.5 ℃, shaking at low speed of 50 r/min for 24 hours, removing the fixing solution, washing with normal saline for 3 times, adding the fixing solution, keeping the temperature constant at 36.5 ℃, shaking at low speed of 50 r/min for 6 hours, washing with normal saline for 3 times, adding cell storage solution, and storing at 2-8 ℃ until the final concentration is 50-500 pieces/. mu.L.
S105, culturing, centrifuging and washing the obtained lactobacillus, and fixing and storing the diluted lactobacillus suspension to obtain the lactobacillus quality control substance.
Specifically, the method comprises the steps of obtaining lactobacillus, subculturing to the third generation, centrifuging and washing for 3-5 times until microscopic examination is qualified, and diluting with normal saline to obtain lactobacillus suspension, wherein the lactobacillus suspension comprises the following steps:
the lactobacillus is purchased, subcultured, and kept for later use after subculture for 3 generations.
Centrifuging cultured and subcultured lactobacillus, washing with normal saline, centrifuging, repeating for 3-5 times, and performing microscopic examination until lactobacillus is cleaned. Diluting with normal saline to obtain lactobacillus suspension.
Fixing and washing the fixing solution and the lactobacillus suspension in a volume ratio of 6:1 twice at 36.5 ℃ at 50 r/min; adding a preservation solution into the lactobacillus suspension after fixation, and preserving at the temperature of 2-8 ℃ to obtain the lactobacillus quality control substance
And (3) fixing the lactobacillus: adding lactobacillus suspension into 6 times volume of cell fixing solution, keeping constant temperature at 36.5 deg.C, shaking at 50 r/min for 24 hr, removing fixing solution, washing with normal saline for 3 times, adding fixing solution, keeping constant temperature at 36.5 deg.C, shaking at 50 r/min for 6 hr, washing with normal saline for 3 times, adding cell storage solution to final concentration of 50-500 pieces/. mu.L, and storing at 2-8 deg.C for use.
The invention provides a quality control substance for detecting visible components of vaginal secretion, and the preparation method of the quality control substance for detecting the visible components of vaginal secretion is applied to the quality control substance for detecting the visible components of vaginal secretion, and the quality control substance for detecting the visible components of vaginal secretion comprises a red blood cell quality control substance, a white blood cell quality control substance, an oral epithelial cell quality control substance, a candida albicans quality control substance and a lactobacillus quality control substance.
In this embodiment, the quality control material for detecting visible components in vaginal secretion further comprises a hemolytic agent, a fixing solution and a preservation solution, wherein the hemolytic agent is a mixed solution of 1-2g/L sodium citrate, 0.3-2.7% triton-100 and 7% NaCl, and the cell preservation solution comprises the following components: the osmotic pressure of the cell preservation solution is 300 +/-40 mOsm/kg, the formaldehyde is 2.5%, 0.05% P300, 8g/L bovine serum albumin, 0.07% sodium chloride, 3.9-7.8 mmol/L glucose, 3.2% -3.8% sodium citrate and the like are prepared as basic formulas. The fixative solution used in this patent comprises the following components: 1.5 to 3.5 percent of formaldehyde. Except red blood cells and white blood cells, the vaginal secretion quality control substance is further enriched by including epithelial cells and lactobacillus quality control substances. The quality control substance of the vaginal secretion, which is formed by red blood cells, white blood cells, epithelial cells and lactobacillus, plays an important role in the quality control of the detection system and the accuracy of the detection result.
According to the invention, the quality control morphology is kept intact on wet microscopic examination, as shown in fig. 2 to 5, wherein fig. 2 is red blood cells, fig. 3 is fungi and epithelial cells (the fungi have small morphology and the epithelial cells have large morphology), fig. 4 is white blood cells and epithelial cells (the white blood cells have small morphology and the epithelial cells have large morphology), and fig. 5 is fungi spores, blastospores and hyphae (the fungi spores basically exist singly and are not propagated; the blastospores are often doubled or piled in the propagation period, and the hyphae are long-strip-shaped when the fungi develop to the most vigorous period); on dry staining microscopy, the nucleus and cytoplasm are clearly distinguished, and the visible components of vaginal secretion can be clearly distinguished, as shown in fig. 6 to 9, wherein fig. 6 is epithelial cells, fig. 7 is white blood cells and red blood cells (small red blood cells, light blue color, large white blood cells, dark blue color), fig. 8 is fungal spores and blastospores (fungal spores exist basically singly and do not propagate; blastospores, propagation stage, often appear in pairs or piles), and fig. 9 is lactobacillus. Can meet the quality control of wet and dry dyeing microscopic examination of visible components of gynecological secretion.
Advantageous effects
1. The problems of cell aggregation, cell expansion, cell shape shrinkage and the like of the tangible quality control substance are solved, and the product quality of the quality control substance is greatly improved.
2. The patent not only comprises quality control substances of red blood cells and white blood cells, but also comprises quality control substances of epithelial cells, candida albicans and lactobacillus, and the types of the quality control substances are expanded.
The quality control material for detecting the visible components of the vaginal secretion comprises a red blood cell quality control material, a white blood cell quality control material, an oral epithelial cell quality control material, a candida albicans quality control material and a lactobacillus quality control material, and after the obtained red blood cells are subjected to anticoagulation whole blood and are separated and purified, the red blood cells are solidified and stored to obtain the red blood cell quality control material; separating and purifying the leucocytes after anticoagulation of the whole blood, and fixing and storing the qualified cell-coated suspension subjected to microscopic examination to obtain the leucocyte quality control substance; subculturing the obtained oral epithelial cells, centrifugally diluting, and fixing and storing an epithelial cell suspension obtained by dilution to obtain the oral epithelial cell quality control substance; culturing, centrifuging and microscopic examining the obtained candida albicans, and fixing and storing a diluted candida albicans suspension to obtain the candida albicans quality control substance; the obtained lactobacillus is cultured, centrifuged and washed, and the diluted lactobacillus suspension is fixed and stored to obtain the lactobacillus quality control substance, except red blood cells and white blood cells, epithelial cells and the lactobacillus quality control substance are included, so that the variety of the quality control substance is enlarged, the product quality is greatly improved, and the accuracy of detecting visible components of vaginal secretion is improved.
While the invention has been described with reference to a preferred embodiment, it will be understood by those skilled in the art that various changes in form and detail may be made therein without departing from the spirit and scope of the invention as defined by the appended claims.
Claims (8)
1. A preparation method of a quality control material for detecting visible components of vaginal secretion is characterized in that the quality control material for detecting the visible components of the vaginal secretion comprises a red blood cell quality control material, a white blood cell quality control material, an oral epithelial cell quality control material, a Candida albicans quality control material and a lactobacillus quality control material, and comprises the following steps:
performing anticoagulation on the obtained red blood cells, separating and purifying, and then performing solidification storage on the red blood cells to obtain the red blood cell quality control substance;
separating and purifying the leucocytes after anticoagulation of the whole blood, and fixing and storing the qualified cell-coated suspension subjected to microscopic examination to obtain the leucocyte quality control substance;
subculturing the obtained oral epithelial cells, centrifugally diluting, and fixing and storing an epithelial cell suspension obtained by dilution to obtain the oral epithelial cell quality control substance;
culturing, centrifuging and microscopic examining the obtained candida albicans, and fixing and storing a diluted candida albicans suspension to obtain the candida albicans quality control substance;
and culturing, centrifuging and washing the obtained lactobacillus, and fixing and storing the diluted lactobacillus suspension to obtain the lactobacillus quality control substance.
2. The method for preparing a quality control substance for detecting the visible components in vaginal secretion according to claim 1, wherein the step of obtaining the quality control substance for red blood cells by performing anticoagulation of whole blood, separation and purification on the whole blood, and then performing solidification and preservation on the red blood cells comprises the steps of:
adding a hemolytic agent into the obtained erythrocyte sample to perform anticoagulation whole blood for 24 hours, centrifuging the anticoagulation whole blood, and diluting and washing the obtained erythrocyte layer for 3-6 times until microscopic examination is qualified to obtain erythrocyte suspension;
and solidifying and storing the erythrocyte suspension to obtain the erythrocyte quality control substance.
3. The method for preparing a quality control substance for detecting the visible components of vaginal secretion according to claim 2, wherein the erythrocyte suspension is solidified and preserved to obtain the erythrocyte quality control substance, and the method comprises the following steps:
fixing the fixing solution and the erythrocyte suspension in a volume ratio of 6:1 for 24 hours at the temperature of 36.5 ℃ and at the speed of 50 r/min, and washing the erythrocyte suspension without the fixing solution with physiological saline for three times to finish the first fixing;
and (3) obtaining the fixing solution again, fixing and washing again according to the method for the first fixing, adding a preservation solution into the erythrocyte suspension after the fixing is finished, and preserving at the temperature of 2-8 ℃.
4. The method for preparing a quality control substance for detecting the visible components of the vaginal secretion according to claim 1, wherein the quality control substance for the oral epithelial cells is obtained by subculturing the obtained oral epithelial cells, centrifugally diluting the oral epithelial cells, and fixing and storing the diluted epithelial cell suspension, and comprises the following steps:
obtaining oral epithelial cells, subculturing to the third generation, centrifuging and washing for 3-5 times until microscopic examination is qualified, and diluting with normal saline to obtain epithelial cell suspension;
and (3) fixing and storing the epithelial cell suspension to obtain the oral epithelial cell quality control substance.
5. The method of claim 4, wherein the step of fixing and preserving the epithelial cell suspension to obtain the oral epithelial cell quality control material comprises:
fixing and washing the epithelial cell suspension and the fixing solution with the volume ratio of 6:1 for two times at 36.5 ℃ under the condition of 50 r/min;
and adding a preservation solution into the epithelial cell suspension after fixation is completed, and preserving at the temperature of 2-8 ℃.
6. The method of claim 5, wherein the fixing solution and the epithelial cell suspension at a volume ratio of 6:1 are fixed and washed twice at 36.5 ℃ and 50 rpm, and the method comprises:
fixing the fixing solution and the erythrocyte suspension in a volume ratio of 6:1 for 24 hours at the temperature of 36.5 ℃ and at the speed of 50 r/min, and washing the epithelial cell suspension without the fixing solution by using physiological saline for three times to finish the first fixing;
re-obtaining the fixing solution, and fixing the fixing solution and the epithelial cell suspension after the first fixing for 6 hours at the temperature of 36.5 ℃ and at the speed of 50 r/min according to the volume ratio of 6: 1;
and (3) washing the epithelial cell suspension after the fixing solution is removed by using physiological saline three times to finish the second fixing of the epithelial cell suspension.
7. A quality control material for detecting the visible component of vaginal secretion, the preparation method of the quality control material for detecting the visible component of vaginal secretion as claimed in any one of claim 1 to claim 6 is applied to the quality control material for detecting the visible component of vaginal secretion,
the quality control substances for detecting visible components in vaginal secretion comprise red blood cell quality control substances, white blood cell quality control substances, oral epithelial cell quality control substances, candida albicans quality control substances and lactobacillus quality control substances.
8. A quality control material for detecting the visible component of vaginal discharge as claimed in claim 7,
the quality control material for detecting the visible components of the vaginal secretion also comprises a hemolytic agent, a stationary liquid and a preservation liquid, wherein the hemolytic agent is a mixed solution of 1-2g/L sodium citrate, 0.3-2.7% triton-100 and 7% NaCl.
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