CN1434127A - Staining kit and preparation method, staining method, and use thereof - Google Patents
Staining kit and preparation method, staining method, and use thereof Download PDFInfo
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- CN1434127A CN1434127A CN 02102739 CN02102739A CN1434127A CN 1434127 A CN1434127 A CN 1434127A CN 02102739 CN02102739 CN 02102739 CN 02102739 A CN02102739 A CN 02102739A CN 1434127 A CN1434127 A CN 1434127A
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Abstract
The present invention relates to a staining kit and its preparation method, staining method and its application in several detection items for detecting canceration cell, red blood cell, campylobacteriosis, vaginal cleaning degree, estrogen level, garlandosus, trichomoniasis, mycosis, diplococcus gonorrhoeae and chlamydi trachomatis on a cervical smear. Said invented kit is formed from fixatino fluid, staining solution and differentiatino solution. Its staining method includes the steps of fixation, staining, differentiation and water-washing. Its film-making speed is quick, only has need of two main, and its detection accuracy rate is high, can be up to 98%.
Description
(1) technical field:
The present invention relates to a kind of staining kit and preparation method thereof, dyeing process and in the quick multinomial pathogenic agent of genital tract infection/sexually transmitted disease (STD) and the application in the cytolgical examination.
(2) background technology:
Traditional dyeing process is Pasteur and HE, belongs to world-renowned cytopathology staining, and its set time is long, and dyeing time is long, and staining procedure is many, and report speed is slow, and the staff is many, and it is few that day is read the sheet amount, the cost height, and complicated operation takes people, time-consuming, expensive.
Ji's nurse Sa (Giemsa) fast staining.Bryon-PA (Patnol Biol 9aris 1996; 24 (1): 75-9) once reported the Giemsa fast staining is used for examination of bone marrow smear.Thatcher-RW in 1977 etc. are used for a conjunctivo-cytology inspection, Howell-WM (HumGenet.1978 in 1978 again; 43 (1): 53-6) be used for the cell chromosome inspection.In the same year, domestic scholars gold tree chastity etc. is used for quick cytodiagnosis malignant tumour, and the author also was used for quick tumour cell diagnosis in 1981.Its dyeing kinetics is fast, only needs 3 minutes, and pair cell nuclear is painted better, but pair cell slurry and cytolemma and neutrophil granule wherein then dye relatively poor.
The blue alcohol liquid of first-class amine fast staining, domestic scholars dragon clean (new medical science, 1978 in 1978; (8): 416) once dyed the tumor tissues printingout, tumour in the operation is diagnosed with this method; My (Chinese journal of medical examination, 1980 of Tang Li in 1980; 3 (1): 42) also be used for cytolgical examination with this liquid.Its dyeing only need 3 minutes, but color separation is relatively poor fast, and cellularstructure is clear inadequately.
The Diff-Quik fast staining, Kass LG (NEWYork:IGAkusHOIN, 1984 in 1984; 15:46) be used to the plate coating checking that punctures.Tollerud DJ{chest in 1989,1989; 95 (37:494-7) } be used for AIDS VICTIMS's lung bundle cysticercus and identify.Its dyeing only needs 2 minutes, but costs an arm and a leg fast.
Dyed blended (" haematol " Wang Fengji chief editor of Rui Shi-Ji's nurse Sa, Tianjin science tech publishing house, nineteen ninety publishes) being mainly used in the blood smear inspection, dyeing process is: select Rui Shi liquid dyeing 25 minutes for use, remove Wright's stain, again with Ji's nurse Sa dye liquor dyeing 4 minutes.After the tap water flushing, put dry voluntarily getting final product in the air.Wright's stain is painted good to endochylema.Ji's nurse Sa liquid is then painted good to examining.This method dyeing kinetics is slower, needs with more than 30 minutes.
To non-gonococcal urinary tract infection, traditional isolated culture is consuming time long, and the microimmunofluorescence method is very stdn not, and is international by SyV2 reagent costliness, all is difficult for promoting
(3) summary of the invention:
The object of the present invention is to provide a kind of rapid dyeing poly functional reagent box.
Another object of the present invention is to provide a kind of preparation method of test kit of the present invention.
Further purpose of the present invention is to provide a kind of dyeing process of test kit of the present invention.
Another object of the present invention is to provide the mentioned reagent box on a cervical smear to the application in the detection of a plurality of projects such as cancerous tumor cell, red blood corpuscle, crooked viable bacteria, vagina cleanness degree, estrogen level, Gardner Salmonella, trichomonad, mould, gonorrhea diplococcus, chlamydia trachomatis.Adapting to examining soon soon controlling of genital tract infection/sexually transmitted disease (STD), and the eager problem of " the genital tract infection intervention engineering " of the birth control of the big face width of solution amount, to realize that cancer knurl patient is early examined the technical object of early controlling.
WG method of the present invention, the abbreviation of the method after being about to Rui Shi (Wright) method and Ji's nurse Sa (Giemsa) fast staining combining.
Staining kit of the present invention is made up of stationary liquid, staining fluid, differentiation liquid,
Stationary liquid wherein: contain 50% anhydrous methanol and 50% dehydrated alcohol (volume percent),
Staining fluid: contain Rui Shi powder 0.5-1.5g, Ji's nurse Sa Shi powder 0.5-1.5g, azure II 0.2-0.6g, glycerine 33-100ml, methyl alcohol (AR) 600-900ml.
Differentiation liquid: anhydrous potassium dihydrogenphosphate 6-7g, disodium hydrogen phosphate,anhydrous 2.5-3g, distilled water 1000ml.
The preparation method of staining kit of the present invention is:
The preparation stationary liquid: will can use after 50% anhydrous methanol and the addition of 50% dehydrated alcohol (volume percent) equivalent,
Preparation staining fluid: be put in 0.5-1.5g Rui Shi powder and 0.5-1.5g Ji's nurse Sa Shi powder in the clean mortar together, after the adding small amount of methanol is fully ground, draw upper strata liquid, add small amount of methanol again, continue to grind, suct a layer liquid again, so continuously for several times, stain fully grinds and dissolves and washes away only in mortar, to going in the Brown Glass Brown glass bottles and jars only, add azure II 0.2-0.6g, glycerine 33-100ml supplies methyl alcohol to 600-900ml.Fully shake once every day, in totally one week, deposits all after-filtration again, stores standby with brown ground glass stoppered bottle.
Preparation differentiation liquid: anhydrous potassium dihydrogenphosphate 6-7g, disodium hydrogen phosphate,anhydrous 2.5-3g, adding distil water 1000ml, adjusting pH value with phosphoric acid salt is that PH6.4-6.8 gets final product.
The dyeing process of staining kit of the present invention the steps include:
Fixing: drip stationary liquid 7-10 and drip on cervical smear, preferred 4-10 drips; Fixing 21-30 second, preferred 15-30 second; The moving slide of side removes stationary liquid.
Dyeing: drip staining fluid 1-4 and drip, preferred 2-3 drips; With dye liquor bedding sample, dye 5-8 second.
Differentiation: drip differentiation liquid 4-14 immediately and drip, preferred 8-12 drips; Rotate slide gently, make it to be mixed, break up 30-38 second with staining fluid.
Washing: slowly wash away raffinate and throw out thereof with tiny flowing water, wipe slide back side moisture, wet sheet microscopy.
Pathocytology and pathogen diagnosis are valued for accurately fast, and non-this is difficult to make reproduction infection and cancer knurl patient to obtain early to examine early to control, and the key problem in technology of quick diagnosis is that a dyeing process fast will be arranged.One minute staining of WGRS of the present invention, the medical detecting method that reached quick, accurate, simple, cheap, inspection item is many, it checks that with two emphasis (cancerous tumor cell+pathogenic agent) combines, form the generaI investigation but also can be used for " fast ten of the cervical smears " that infectious diseases in genital tract is checked of not only being used to give protection against cancer, promptly can be on a cervical smear to the detection of a plurality of projects such as cancerous tumor cell, red blood corpuscle, crooked viable bacteria, vagina cleanness degree, estrogen level, Gardner Salmonella, trichomonad, mould, gonorrhea diplococcus, chlamydia trachomatis.
To malignant cell morphological feature under the WG dyeing, from the cytopathology angle, various tumour cells are carried out the fast qualitative diagnosis, verify through the clinical examination of 13346 on-the-spot examples of Operation theatre, Endoscopy Room, anti-cancer and sexually transmitted disease (STD) generaI investigation.
To infectious diseases in genital tract, under WG dyeing, can know the morphological specificity of the various pathogenic agent of demonstration genital tract infection/sexually transmitted disease (STD), show by 12205 routine papanicolaou test results, can quick diagnosis cause the multiple disease of genital tract infection.As bacterial vaginitis, gonococcal vaginitis, non-gonococcal vaginitis, trichomonal vaginitis, colpitis mycotica etc.
Following table is with pap staining 12531 examples, HE 13746 examples that dye, and the cervical smear of WG rapid dyeing 10185 examples is done contrast and is observed, and its comparing result is shown in subordinate list:
| Item compared | Pasteur's (simplification) | HE (simplification) | WG is quick |
| Set time | 15 minutes | 15 minutes | Half a minute |
| Dyeing time | 60 minutes | 30 minutes | Half a minute |
| Staining procedure | 14 steps, microscopy after the sealing | 7 steps are with a left side | 3 steps, not sealing, wet sheet microscopy |
| Report speed | About half a day | About half a day | About 2 minutes |
| The staff | ????2 | ????2 | ????1 |
| Day is read the sheet amount | About 50 examples | About 50 examples | About 100 examples |
| The cost ratio | ????20： | ????10： | ????1 |
Passed through checking and verifying of 30,000 many cases, prove that the outstanding advantage of this method is: 1. film-making speed is fast, only needs two minutes; 2. accuracy rate height, full accuracy rate of diagnosis can reach 98%; 3. the equipment simple operations is easy, can be at the Operation theatre scene, and the Endoscopy Room scene, carry out at anti-cancer and sexually transmitted disease (STD) generaI investigation scene; 4. expense is cheap, is fit to national conditions, is convenient to promote; 5. Color is good, can clearly illustrate the cell level of uterus vagina epithelium, the influence degree of cell rank and estrogen level, also fixedly dewatering agent is few because of using, and the cell that seems is bigger, stretch good, natural shape, distortion is slight, the tumour cell of especially differentiation difference, malignant characteristics is outstanding, is easy to diagnosis.
Generally apply along with of the present invention, will play a role in promoting for the enforcement of " the genital tract infection intervention engineering " of the numerous women of child-bearing age's healthy reproduction and the big face width of adaptive capacity.
(5) embodiment:
The following examples can make those skilled in the art more fully understand the present invention, but do not limit the present invention in any way.
Embodiment 1:
Get after 50% anhydrous methanol and the addition of 50% dehydrated alcohol (volume percent) equivalent as stationary liquid;
With Rui Shi powder 1.5g and Ji's nurse Sa Shi powder 1.5g, with being put in the clean mortar, after the adding small amount of methanol is fully ground, draw upper strata liquid, add small amount of methanol again, continue to grind, suct a layer liquid again, so continuously for several times, stain fully grinds and dissolves and washes away only in mortar, to going in the Brown Glass Brown glass bottles and jars only, adds azure II 0.6g, glycerine 100ml, supply methyl alcohol to 900ml as staining fluid.Fully shake once every day, in totally one week, deposits all after-filtration again, stores standby with brown ground glass stoppered bottle.
Get anhydrous potassium dihydrogenphosphate 7g, disodium hydrogen phosphate,anhydrous 3g, adding distil water 1000ml is as differentiation liquid, and adjusting pH value with phosphoric acid salt is that PH6.8 gets final product.
Test kit is formed (50 person-portion), one bottle of stationary liquid 20ML, one bottle of staining fluid 10ML, two bottles of differentiation liquid 20ML.
Embodiment 2:
Drip 6 of stationary liquids on cervical smear, fixed for 20 seconds, the moving slide of side removes stationary liquid.Drip 2 of staining fluids,, dyed for 5 seconds dye liquor bedding sample.Drip 8 of differentiation liquid immediately, rotate slide gently, make it to be mixed, broke up for 38 seconds with staining fluid.Slowly wash away raffinate and throw out thereof with tiny flowing water, wipe slide back side moisture, wet sheet microscopy.
Attention: 1. with before must bottleneck is logical with pinprick so that dye liquor can unimpeded oozing.2. when scraping (being coated with) sheet, want suitably the position, and specimen amount is wanted foot, and it is light that gimmick is wanted, and smears and will approach.The very few major cause of failing to pinpoint a disease in diagnosis often of sample.3. dyeing time can be according to temperature, the sample thin and thick, and proper extension, but can not shorten.4. test kit should be put shady and cool place, preferably preserves validity period 3-4 in 2-8 ℃ of refrigerator.
Staining reaction: epithelial cell is red-purple, and endochylema is pale blue, and white corpuscle nuclear is red-purple, and basophilic stippling is hyacinthine in the slurry, and the acidophilia particle is orange red, and neutrophilic granule is redly little, and red blood corpuscle is orange red.Trichomonad is a pale blue, is incarnadine in the shuttle nuclear of head.Fungal hyphae is a dark-brown, the gemma same mycelia of developing the color.Chlamydozoan is yellowish to dark-brown.Diplococcus and Jia Dena Salmonella are light gray.
Claims (5)
1. a staining kit is made up of stationary liquid, staining fluid, differentiation liquid,
Stationary liquid wherein: contain 50% anhydrous methanol and 50% dehydrated alcohol (volume percent),
Staining fluid: contain Rui Shi powder 0.5-1.5g, Ji's nurse Sa Shi powder 0.5-1.5g, azure II 0.2-0.6g, glycerine 33-100ml, methyl alcohol (AR) 600-900ml.
Differentiation liquid: anhydrous potassium dihydrogenphosphate 6-7g, disodium hydrogen phosphate,anhydrous 2.5-3g, distilled water 1000ml.
2. the preparation method of claim 1 staining kit is:
The preparation stationary liquid: will can use after 50% anhydrous methanol and the addition of 50% dehydrated alcohol (volume percent) equivalent,
Preparation staining fluid: be put in 0.5-1.5g Rui Shi powder and 0.5-1.5g Ji's nurse Sa Shi powder in the clean mortar together, after the adding small amount of methanol is fully ground, draw upper strata liquid, add small amount of methanol again, continue to grind, suct a layer liquid again, so continuously for several times, stain fully grinds and dissolves and washes away only in mortar, to going in the Brown Glass Brown glass bottles and jars only, add azure II 0.2-0.6g, glycerine 33-100ml supplies methyl alcohol to 600-900ml.Fully shake once every day, in totally one week, deposits all after-filtration again, stores standby with brown ground glass stoppered bottle.
Preparation differentiation liquid: anhydrous potassium dihydrogenphosphate 6-7g, disodium hydrogen phosphate,anhydrous 2.5-3g, adding distil water 1000mL, adjusting pH value with phosphoric acid salt is that PH6.4-6.8 gets final product.
3. the dyeing process of the staining kit of claim 1 the steps include:
Fixing: drip a stationary liquid 7-10 and drip on cervical smear, fixedly 21-30 second, side is moved slide and is removed stationary liquid.
Dyeing: drip staining fluid 1-4 and drip,, dye 5-8 second with dye liquor bedding sample.
Differentiation: drip differentiation liquid 4-14 immediately and drip, rotate slide gently, make it to be mixed, break up 30-38 second with staining fluid.
Washing: slowly wash away raffinate and throw out thereof with tiny flowing water, wipe slide back side moisture, wet sheet microscopy.
4. dyeing process according to claim 3 the steps include:
Fixing: drip a stationary liquid 4-10 and drip on cervical smear, fixedly 15-30 second, side is moved slide and is removed stationary liquid.
Dyeing: drip staining fluid 2-3 and drip,, dye 5-8 second with dye liquor bedding sample.
Differentiation: drip differentiation liquid 8-12 immediately and drip, rotate slide gently, make it to be mixed, break up 30-38 second with staining fluid.
Washing: slowly wash away raffinate and throw out thereof with tiny flowing water, wipe slide back side moisture, wet sheet microscopy.
The staining kit of claim 1 on a cervical smear to the application in the detection of a plurality of projects such as cancerous tumor cell, red blood corpuscle, crooked viable bacteria, vagina cleanness degree, estrogen level, Gardner Salmonella, trichomonad, mould, gonorrhea diplococcus, chlamydia trachomatis.
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| CN1167807C CN1167807C (en) | 2004-09-22 |
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| CN103792126A (en) * | 2013-11-20 | 2014-05-14 | 重庆市农业科学院 | Staining solution applicable to tea geometrid blood cells and staining method thereof |
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| CN104472331A (en) * | 2014-12-01 | 2015-04-01 | 西北农林科技大学 | Fast transparentizing method for observing fungi of plant roots |
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| CN103792126B (en) * | 2013-11-20 | 2016-06-15 | 重庆市农业科学院 | The staining fluid of tea geometrid hemocyte and dyeing process thereof |
| CN103792126A (en) * | 2013-11-20 | 2014-05-14 | 重庆市农业科学院 | Staining solution applicable to tea geometrid blood cells and staining method thereof |
| CN103808550A (en) * | 2014-02-19 | 2014-05-21 | 上海海洋大学 | Myxosporean dyeing and sealed-storage method convenient for morphological observation |
| CN103808550B (en) * | 2014-02-19 | 2016-08-17 | 上海海洋大学 | A kind of Myxosporean dyeing method of seal being easy to morphological observation |
| CN104472331A (en) * | 2014-12-01 | 2015-04-01 | 西北农林科技大学 | Fast transparentizing method for observing fungi of plant roots |
| CN104472331B (en) * | 2014-12-01 | 2016-11-23 | 西北农林科技大学 | Fast transparent method for observation of plant root system fungus |
| CN105067412A (en) * | 2015-08-10 | 2015-11-18 | 长春瑞克医疗科技有限公司 | Vaginal secretion staining fluid, and preparation method and staining method thereof |
| CN105067412B (en) * | 2015-08-10 | 2018-06-26 | 迪瑞医疗科技股份有限公司 | A kind of vaginal fluid dyeing liquor and preparation method thereof and colouring method |
| CN105300772A (en) * | 2015-09-30 | 2016-02-03 | 成都华西海圻医药科技有限公司 | Wright-Giemsa compound staining solution and preparation method thereof |
| CN107664595A (en) * | 2016-07-27 | 2018-02-06 | 石家庄晟科生物科技有限公司 | Efficient chromosomal G-banding staining kit |
| CN107300495A (en) * | 2017-06-15 | 2017-10-27 | 托克逊县夏乡畜牧业服务中心 | A kind of colouring method of pyriform worm blood film compound stain solution and blood film |
| CN107589117A (en) * | 2017-08-22 | 2018-01-16 | 贵州省畜牧兽医研究所 | A kind of quick microscopy diagnostic kit of swine eperythrozoonosis and preparation method thereof |
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