CN109115584A - The detection reagent and its application of Hemoglobin F in a kind of detection red blood cell - Google Patents

The detection reagent and its application of Hemoglobin F in a kind of detection red blood cell Download PDF

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Publication number
CN109115584A
CN109115584A CN201811064880.3A CN201811064880A CN109115584A CN 109115584 A CN109115584 A CN 109115584A CN 201811064880 A CN201811064880 A CN 201811064880A CN 109115584 A CN109115584 A CN 109115584A
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China
Prior art keywords
detection reagent
solid
red blood
hemoglobin
distilled water
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Pending
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CN201811064880.3A
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Chinese (zh)
Inventor
朱宏远
樊斐
翟燕红
曹正
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BEIJING OBSTETRICS AND GYNECOLOGY HOSPITAL CAPITAL MEDICAL UNIVERSITY
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BEIJING OBSTETRICS AND GYNECOLOGY HOSPITAL CAPITAL MEDICAL UNIVERSITY
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Priority to CN201811064880.3A priority Critical patent/CN109115584A/en
Publication of CN109115584A publication Critical patent/CN109115584A/en
Pending legal-status Critical Current

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    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00Sampling; Preparing specimens for investigation
    • G01N1/28Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/30Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10Investigating individual particles
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00Sampling; Preparing specimens for investigation
    • G01N1/28Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/30Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
    • G01N2001/302Stain compositions
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00Sampling; Preparing specimens for investigation
    • G01N1/28Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/30Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
    • G01N2001/305Fixative compositions
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/01Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials specially adapted for biological cells, e.g. blood cells
    • G01N2015/012Red blood cells
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10Investigating individual particles
    • G01N2015/1006Investigating individual particles for cytology
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
    • G01N15/10Investigating individual particles
    • G01N2015/1024Counting particles by non-optical means

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  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Dispersion Chemistry (AREA)
  • Molecular Biology (AREA)
  • Biomedical Technology (AREA)
  • Engineering & Computer Science (AREA)
  • Investigating Or Analysing Biological Materials (AREA)

Abstract

The present invention relates to the applications of the detection reagent and the detection reagent of Hemoglobin F in a kind of detection red blood cell, the detection reagent includes acidic effluent liquid and dyeing liquor, and the acidic effluent liquid is prepared by following raw material: 25% hydrochloric acid solution, hematoxylin solid, ferric trichloride solid, 95% ethanol solution, distilled water;The dyeing liquor is prepared by following raw material: Yihong solid, distilled water.Detection reagent preparation is simple, and recall rate is high.

Description

The detection reagent and its application of Hemoglobin F in a kind of detection red blood cell
Technical field
The present invention relates to a kind of detection reagent, in particular in a kind of detection red blood cell the detection reagent of Hemoglobin F with And the application of the detection reagent.
Background technique
In recent years, tire mother transfusion syndrome gradually caused the attention of ob-gyn, how to early diagnose out this hair Emphasis disease concealment and the disease of serious consequence is easily caused to pay close attention to jointly as ob-gyn and clinical laboratory doctor.Due to normal 99.9% red blood cell contains hemoglobin A without Hemoglobin F in adult human body, and red blood cell contains blood red egg in fetus body White F does not contain hemoglobin A, therefore can be by Hemoglobin F content in detection pregnant and lying-in women's body, between judging whether that generation tire is female The case where blood transfusion.Known Hemoglobin F has stronger capacity antacid than hemoglobin A.Therefore, the Blood piece after fixation, sets Held for some time in acidic buffer, the only red blood cell containing Hemoglobin F are not eluted, such as with eosin stains in scarlet Color.Hemoglobin F acid elution test (K-B experiment) is fetus in the most common quantitative detection maternal blood circulation of current each hospital Red blood cell number purpose method.Hemoglobin is carried out according to the method for " national clinical examination operating instruction " (third edition) defined F acid elution test finds influence of this method result vulnerable to external factors such as temperature, manual operation factors, and when testing Between it is longer.And this law is ineffective for the red blood cell elution without containing Hemoglobin F, causes to judge whether red blood cell contains The differentiation of Hemoglobin F result is unobvious, and as a result accuracy is poor.
Summary of the invention
The purpose of the present invention is to provide a kind of detection reagents of Hemoglobin F in new detection red blood cell, on solving State the problem of proposing in background technique.
To achieve the above object, the invention provides the following technical scheme:
The detection reagent of Hemoglobin F in a kind of detection red blood cell, including acidic effluent liquid and dyeing liquor;The acidity is washed De- liquid is prepared by following raw material: 25% hydrochloric acid solution, hematoxylin solid, ferric trichloride solid, 95% ethanol solution, distilled water; The dyeing liquor is prepared by following raw material: Yihong solid, distilled water;The acidic effluent liquid the preparation method is as follows: by 0.75 Gram hematoxylin solid is completely dissolved in 100 milliliter of 95% ethanol solution, mixes, and mixture A is made;2.4 grams of ferric trichlorides are consolidated Body is completely dissolved in 2 milliliter of 25% hydrochloric acid solution, and distilled water is added later to 100 milliliters, mixes, mixture B is made;Mixture A and mixture B is uniformly mixed according to 5: 1 ratio of volume ratio, obtains the acidic effluent liquid;The preparation method of the dyeing liquor It is as follows: 1 gram of Yihong solid being completely dissolved in 100 milliliters of distilled water, mixes, obtains the dyeing liquor.
A kind of application of detection reagent as described above, its step are as follows: (1) Blood piece with 80% ethyl alcohol is fixed 5 minute, The Blood piece fixed is placed in 40 DEG C -50 DEG C of the acidic effluent liquid and keeps the temperature 1 minute, washed to dry;It (2) will be after drying Blood piece is dyed 1 minute with the dyeing liquor, is washed to dry;(3) Blood piece after drying is placed under microscope, is knitted referring to net red thin Born of the same parents' method of counting counts percentage shared by Hemoglobin F positive red blood cell in 1000 red blood cells.
A kind of purposes of detection reagent as described above, the detection reagent be used to detect Hemoglobin F in red blood cell In shared percentage.
Compared with prior art, the beneficial effects of the present invention are: detection reagent of the invention is to without containing Hemoglobin F Red blood cell eluting power is stronger, so that whether there is or not Hemoglobin F fruiting areas to show clearly for red blood cell, improves observer and is easy visual fatigue Situation, convenient for counting under microscope, count results accuracy is good.Also, Hemoglobin F is carried out with detection reagent of the invention Acid elution test is substantially shorter test period, reduces operating procedure, while reducing extraneous factor probability of interference, convenient for experiment Room quickly checks.
Detailed description of the invention
In order to illustrate more clearly of technical solution of the present invention, the present invention is made with reference to the accompanying drawings and examples further Explanation.
Attached drawing 1 show the colored graph for detecting Hemoglobin F in maternal blood using the detection reagent of embodiment 1 Piece;
Attached drawing 2 show the colored graph for detecting Hemoglobin F in maternal blood using the detection reagent of embodiment 2 Piece.
Specific embodiment
Below in conjunction with embodiment, to technical solution of the present invention carry out it is clear, be fully described by.Based in the present invention Embodiment, every other embodiment obtained by those of ordinary skill in the art without making creative efforts, It shall fall within the protection scope of the present invention.
Embodiment 1
In the present embodiment, a kind of detection reagent detecting Hemoglobin F in red blood cell, including acidic effluent liquid and dyeing Liquid;The acidic effluent liquid is prepared by following raw material: 25% hydrochloric acid solution, hematoxylin solid, ferric trichloride solid, 95% ethyl alcohol Solution, distilled water;The dyeing liquor is prepared by following raw material: Yihong solid, distilled water.
Preparation method includes the following steps:
(1) preparation of mixed liquor A: 0.75 gram of hematoxylin solid is sufficiently dissolved with 100 milliliter of 95% ethanol solution, is mixed It is even;
(2) it the preparation of mixed liquid B: after 2.4 grams of ferric trichloride solids are dissolved in 2 milliliter of 25% hydrochloric acid solution, are added and steam Distilled water is mixed to 100 milliliters;
(3) preparation of acidic effluent liquid: by mixed liquor A and mixed liquid B, 5: 1 ratios are uniformly mixed by volume;Acidity is washed The pH value of de- liquid is 1.65 ± 0.2;
(4) preparation of dyeing liquor: 1 gram of Yihong solid is dissolved in 100 milliliters of distilled water, is mixed.
Detection reagent described in the invention uses step:
(1) Blood piece is fixed 5 minutes with 80% ethyl alcohol, it is accurate the Blood piece fixed to be placed in 40 DEG C of acidic effluent liquid After heat preservation 1 minute, wash to dry;
(2) it after dyeing the Blood piece after drying 1 minute with dyeing liquor, washes to dry;
(3) Blood piece after drying is placed under microscope, counts 1000 red blood cells referring to reticulocyte count method Percentage shared by middle Hemoglobin F positive red blood cell.
Embodiment 2
In the present embodiment, a kind of detection reagent detecting Hemoglobin F in red blood cell, including acidic effluent liquid and dyeing Liquid;The acidic effluent liquid is prepared by following raw material: 25% hydrochloric acid solution, hematoxylin solid, ferric trichloride solid, 95% ethyl alcohol Solution, distilled water;The dyeing liquor is prepared by following raw material: Yihong solid, distilled water.
Preparation method includes the following steps:
(1) preparation of mixed liquor A: 0.75 gram of hematoxylin solid is sufficiently dissolved with 100 milliliter of 95% ethanol solution, is mixed It is even;
(2) it the preparation of mixed liquid B: after 2.4 grams of ferric trichloride solids are dissolved in 2 milliliter of 25% hydrochloric acid solution, are added and steam Distilled water is mixed to 100 milliliters;
(3) preparation of acidic effluent liquid: by mixed liquor A and mixed liquid B, 5: 1 ratios are uniformly mixed by volume;Acidity is washed The pH value of de- liquid is 1.65 ± 0.2;
(4) preparation of dyeing liquor: 1 gram of Yihong solid is dissolved in 100 milliliters of distilled water, is mixed.
Detection reagent described in the invention uses step:
(1) Blood piece is fixed 5 minutes with 80% ethyl alcohol, it is accurate the Blood piece fixed to be placed in 50 DEG C of acidic effluent liquid After heat preservation 1 minute, wash to dry;
(2) it after dyeing the Blood piece after drying 1 minute with dyeing liquor, washes to dry;
(3) Blood piece after drying is placed under microscope, counts 1000 red blood cells referring to reticulocyte count method Percentage shared by middle Hemoglobin F positive red blood cell.
Detection reagent made from above-described embodiment 1-2 is applied to red blood cell of the observation containing Hemoglobin F.
Observe result: the red blood cell containing Hemoglobin F is colored liquid and dyes cerise, is positive cell, does not contain blood The red blood cell of Lactoferrin F only leaves the cell membrane person of light shadow, for negative red blood cell.
It is that Hemoglobin F is detected in maternal blood using the detection reagent in above-described embodiment 1 shown in attached drawing 1 Picture is dyed, wherein dark cell is positive cell.It is to be existed shown in attached drawing 2 using the detection reagent in above-described embodiment 2 The dyeing picture of Hemoglobin F is detected in maternal blood, wherein dark cell is positive cell.
It is obvious to a person skilled in the art that invention is not limited to the details of the above exemplary embodiments, this is fallen in All changes in the meaning and scope of invention the meaning and scope of the equivalent requirements of the claims are included within the present invention.

Claims (3)

1. the detection reagent of Hemoglobin F in a kind of detection red blood cell, including acidic effluent liquid and dyeing liquor;
The acidic effluent liquid is prepared by following raw material: 25% hydrochloric acid solution, hematoxylin solid, ferric trichloride solid, 95% second Alcoholic solution, distilled water;
The dyeing liquor is prepared by following raw material: Yihong solid, distilled water;
The acidic effluent liquid the preparation method is as follows: 0.75 gram of hematoxylin solid is completely dissolved in 100 milliliter of 95% ethyl alcohol Solution mixes, and mixture A is made;2.4 grams of ferric trichloride solids are completely dissolved in 2 milliliter of 25% hydrochloric acid solution, are added later Distilled water is mixed to 100 milliliters, and mixture B is made;Mixture A and mixture B is uniformly mixed according to 5: 1 ratio of volume ratio, i.e., The acidic effluent liquid is made;
The dyeing liquor the preparation method is as follows: 1 gram of Yihong solid is completely dissolved in 100 milliliters of distilled water, mix, that is, make Obtain the dyeing liquor.
2. a kind of application of detection reagent as described in claim 1, which is characterized in that steps are as follows:
(1) Blood piece is fixed 5 minutes with 80% ethyl alcohol, the Blood piece fixed is placed in 40 DEG C -50 DEG C of the acidic effluent liquid Middle heat preservation 1 minute, is washed to dry;
(2) dyeing liquor of the Blood piece after drying is dyed 1 minute, is washed to dry;
(3) Blood piece after drying is placed under microscope, counts blood in 1000 red blood cells referring to reticulocyte count method Percentage shared by Lactoferrin F positive red blood cell.
3. a kind of purposes of detection reagent as described in claim 1, which is characterized in that the detection reagent be used to detect blood Lactoferrin F shared percentage in red blood cell.
CN201811064880.3A 2018-09-10 2018-09-10 The detection reagent and its application of Hemoglobin F in a kind of detection red blood cell Pending CN109115584A (en)

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1234117A (en) * 1996-10-21 1999-11-03 应用成像公司 Use of anti-embryonic hemoglobin antibodies to identify fetal cells
US20030157113A1 (en) * 1999-12-28 2003-08-21 Terman David S. Compositions and methods for treatment of neoplastic disease

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1234117A (en) * 1996-10-21 1999-11-03 应用成像公司 Use of anti-embryonic hemoglobin antibodies to identify fetal cells
US20030157113A1 (en) * 1999-12-28 2003-08-21 Terman David S. Compositions and methods for treatment of neoplastic disease

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
SAMUEL S. ANTWI-BAFFOUR ET AL: "Foetomaternal Haemorrhage (FMH): A Case for Routine Screening", 《JOURNAL OF GYNECOLOGY AND OBSTETRICS,》 *

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