阅读说明：本技术 一种红细胞渗透脆性测定方法及其试剂盒 (A kind of osmotic fragility measuring method and its kit ) 是由 廉正鑫 祁晖 于 2019-09-04 设计创作，主要内容包括：本发明提供一种红细胞渗透脆性测定方法及其试剂盒,其中测定方法包括：S1、在两个比色杯中分别加入等量的不同浓度的低渗透压盐溶液,其中一管是不完全溶血管,其加入的低渗透压盐溶液不能使全血完全溶血,另一管是完全溶血管,其加入的低渗透压盐溶液可以使全血完全溶血；S2、分别往两个比色杯中加入等量全血,混匀后,立即检测不完全溶血管中混合液的散射光值A0、完全溶血管中混合液的散射光值B0；S3、在溶血结束后的某一特定时间,测定不完全溶血管中混合液的的散射光值A1、完全溶血管中混合液的B1；S4、根据公式：溶血百分率%=(A0-A1)/(B0-B1)×100%计算不完全溶血管的溶血百分率。该测定方法的检测结果更精确,且能够实现结果标准化和一致性。(The present invention provides a kind of osmotic fragility measuring method and its kit, wherein measuring method include: S1, be separately added into two cuvettes equivalent various concentration Hyposmolality salting liquid, wherein a pipe is incomplete haemolysis pipe, its Hyposmolality salting liquid being added cannot make whole blood complete hemolysis, another pipe is complete hemolysis pipe, and the Hyposmolality salting liquid being added can make whole blood complete hemolysis；S2, equivalent whole blood is added into two cuvettes respectively, after mixing, detect immediately the scattering light value A0 of mixed liquor in incomplete haemolysis pipe, in complete hemolysis pipe mixed liquor scattering light value B0；S3, a certain specific time after haemolysis, measure the scattering light value A1 of mixed liquor in incomplete haemolysis pipe, in complete hemolysis pipe mixed liquor B1；S4, according to formula: percentage of hemolysis %=(A0-A1)/(B0-B1) × 100% calculates the percentage of hemolysis of incomplete haemolysis pipe.The testing result of the measuring method is more accurate, and can be realized result standardization and consistency.)

1. a kind of osmotic fragility measuring method, which comprises the following steps:

S1, be separately added into two cuvettes equivalent various concentration Hyposmolality salting liquid, wherein a pipe is incomplete Haemolysis pipe, the Hyposmolality salting liquid being added cannot make whole blood complete hemolysis, and another pipe is complete hemolysis pipe, and what is be added is low Osmotic pressure salting liquid can make whole blood complete hemolysis；

S2, equivalent whole blood is added into described two cuvettes respectively, after mixing, detects mixed liquor in incomplete haemolysis pipe immediately Scattering light value A0, in complete hemolysis pipe mixed liquor scattering light value B0；

S3, a certain specific time after haemolysis, measure mixed liquor in incomplete haemolysis pipe scattering light value A1, completely The B1 of mixed liquor in haemolysis pipe；

S4, according to formula: percentage of hemolysis %=(A0-A1)/(B0-B1) × 100% calculates the haemolysis percentage of incomplete haemolysis pipe Rate.

2. osmotic fragility measuring method according to claim 1, which is characterized in that the incomplete haemolysis pipe Hyposmolality salting liquid includes inorganic salts and buffer and at least one of the preservatives.

3. osmotic fragility measuring method according to claim 1, which is characterized in that the complete hemolysis pipe it is low Osmotic pressure salting liquid includes inorganic salts, buffer and at least one of the preservatives.

4. osmotic fragility measuring method according to claim 2 or 3, which is characterized in that the inorganic salts are selected from At least one of sodium chloride, potassium chloride, magnesium chloride, sodium sulphate, magnesium sulfate.

5. osmotic fragility measuring method according to claim 4, which is characterized in that the preservative is selected from At least one of Proclin300 and Sodium azide.

6. osmotic fragility measuring method according to claim 5, which is characterized in that the buffer is selected from phosphoric acid At least one of salt buffer, Glycine-NaOH buffer, Tris- hydrochloride buffer, HEPES buffer solution.

7. osmotic fragility measuring method according to claim 2, which is characterized in that the incomplete haemolysis pipe In Hyposmolality salting liquid, the inorganic salts use concentration for the sodium chloride of 3.5g/L-4.5g/L or osmotic pressure it is comparable other Inorganic salts.

8. osmotic fragility measuring method according to claim 3, which is characterized in that the complete hemolysis pipe it is low The sodium chloride solution or its osmotic pressure and 0.1g/ that osmotic pressure salting liquid is distilled water or pure water or concentration is 0.1g/L-2g/L Other comparable buffer salt solutions of the osmotic pressure of the sodium chloride solution of L-2g/L.

9. osmotic fragility measuring method according to claim 1, which is characterized in that the whole blood and not exclusively it is molten The volume ratio of Hyposmolality salting liquid is 1:50-500 in blood vessel.

10. a kind of osmotic fragility assay kit is suitable for osmotic fragility as described in claim 1 and measures Method characterized by comprising the Hyposmolality salting liquid of incomplete haemolysis pipe and the Hyposmolality salt of complete hemolysis pipe are molten The Hyposmolality salting liquid of liquid, the incomplete haemolysis pipe includes inorganic salts and buffer and at least one of the preservatives, The Hyposmolality salting liquid of the complete hemolysis pipe includes inorganic salts, buffer and at least one of the preservatives.

Technical field

The present invention relates to osmotic fragility determination techniques fields, and in particular to a kind of osmotic fragility measurement side Method and its kit.

Background technique

The ordinary life of mature erythrocyte is about 120 days in human body, about walks 500 public affairs during survival in circulating system In, the blood vessel and hole more much smaller than diameter itself are repeatedly squeezed through, therefore red blood cell must have good toughness and deformation energy Power.Red blood cell maintains morphotropism and stability mainly by erythrocyte membrane, if certain albumen changes on film, can cause molten Blood, such as hereditary spherocytosis.

Osmotic fragility is to examine a clinical common counter of erythrocyte membrane defect, erythrocyte osmotic fragility test It is measurement red blood cell in the hypotonic saline solution of various concentration, the water swelling ability that can bear.Normally red blood cell is Concave-concave disc, if red blood cell is placed in hypotonic solution, because intraor extracellular is there are permeable pressure head, hydrone enters red blood cell, It brings it about swelling or even red blood cell rupture and haemolysis occurs.There is the feature of haemolysis in Hyposmolality salting liquid in red blood cell As osmotic fragility depends primarily on the surface to volume ratio of red blood cell.The big and small in size person couple of surface area The hypotonic easy resistance of salt water is larger (brittleness is smaller), and on the contrary then resistance is smaller (brittleness increase).(Shang Hong, Wang Yusan, Shen The sub- fine jade, national clinical examination operating instruction (the 4th edition) Beijing [M]: People's Health Publisher, 2015.66).

Osmotic fragility increase sees hereditary spherocytosis, hereditary elliptocytosis and part Hereditary stomatocytosis is also found in autoimmune hemolytic anemia.Osmotic fragility reduction sees various pearl egg White dyspoiesis anaemia, HbC, HbD, HbE disease, hypoferric anemia, thalassemia, Postsplenectomy and some other red thin Disease such as liver diseases of after birth exception etc..(Shang Hong, Wang Yusan, Shen Ziyu, national clinical examination operating instruction (the 4th edition) The Beijing [M]: People's Health Publisher, 2015.66)

The method for clinically detecting osmotic fragility at present mainly has following three kinds:

One, traditional-handwork method (Shang Hong, Wang Yusan, Shen Ziyu, national clinical examination operating instruction (the 4th edition) Beijing [M]: the people Hygienic publishing house, 2015.66):

Detecting step is as follows:

(1) the dry small test tube 14 of cleaning is taken, each pipe is by table 1 plus distilled water and 10g/L sodium chloride solution.

(2) examinee venous blood 1mL being taken with dry sterilization syringe, needle slope is upward, and it is flat to hold syringe, pass through syringe needle 1 drop whole blood is added in every pipe, gently shakes up；Normal person's blood is in kind taken to be added on Normal group test tube.

(3) by each pipe rest chamber middle benefit gas 2 hours, whole 14 pipe haemolysises from high concentration.

Test tube number	1	2	3	4	5	6	7	8	9	10	11	12	13	14
															Distilled water (drop)	20	19	18	17	16	15	14	13	12	11	10	9	8	7
10g/LNaCl (drop)	5	6	7	8	9	10	11	12	13	14	15	16	17	18
															Sodium chloride concentration (g/L)	2.0	2.4	2.8	3.2	3.6	4.0	4.4	4.8	5.2	5.6	6.0	6.4	6.8	7.2

1 sodium chloride solution dilution table of table

Two, (Wang Weiwei, Yuan Xiangliang, Fei Qili, Shen Lisong, flow cytometry are quick for improvement Sanford method (simple semiquantitative method) Application [J] laboratory medicine of measurement osmotic fragility, 2016;31 (10), 911-915):

Detecting step is as follows:

13 test tube numbers are taken, the NaCl solution of following concentration: 1.0 g/L, 2.0 g/ is configured to 9g/L NaCl and sterile water L、3.0 g/L、3.5 g/L、4.0 g/L、4.5 g/L、5.0 g/L、5.5 g/L、6.0 g/L、6.5 g/L、7.0 g/L、7.5 g/L,8.5 g/L；

Every pipe adds 50 μ l of anticoagulant heparin whole blood, is gently mixed by inversion, (20 DEG C) placement 30min of room temperature；

Each pipe is mixed 1 time, 3000 × g is centrifuged 3min, takes 200 μ l of supernatant to 96 hole flat undersides, measures each pipe supernatant extinction Spend A value；

With the A value that the A value of 1.0 g/L NaCl complete hemolysis pipes is 100%, You Geguan calculate corresponding percentage of hemolysis (%)= (measurement pipe A value/complete hemolysis pipe A value) × 100%, the NaCl concentration to there is 50% haemolysis judges concentration as erythrocyte fragility；

Result judgement: the NaCl concentration range of normocyte haemolysis is 4.00-4.45g/L, when erythrocyte hemolysis to be measured When NaCl concentration is higher than this range limit, it is determined as that osmotic fragility increases.

Three, a tube method: this method is a kind of method that Du Chuanshu is innovated on the basis of manual method, uses instrument Interpretation is carried out, and is commercially produced and has been sold by Guangzhou meter Ji Medical Devices Co., Ltd., is that Hospitals at Present is wide General receiving and a kind of method used.Specific measurement method is as follows:

(1) the Hyposmolality salting liquid and hemolytic agent of certain certain concentration are prepared.

(2) red blood cell suspension of two parts of equivalent is taken to be respectively placed in the Hyposmolality salting liquid and hemolytic agent of certain certain concentration In, it mixes.

After (3) 37 DEG C of warm bath 5min.

(4) absorbance A and C of two parts of solution are measured respectively using 530nm wavelength spectrophotometer.

(5) hemolysis rate %=(1-A/C) × 100%.

(6) reference value: < 65%.

In addition to this, new or improvement osmotic fragility detection method is proposed there are also some scientific research personnel, But these technologies are not yet universal in clinical hospitals, comprising:

1, (Ye great Tian, Li Lei, Zhou Yahao are red thin for examining for the osmotic fragility method of inspection based on micro-fluidic chip The micro-fluidic chip of born of the same parents' osmotic fragility: China, 101464458B [P] .2013-01-09.):

Detecting step is as follows:

(1) blood sample is inputted in the different detection cells in micro-fluidic chip by blood sample import；

(2) NaCl solution of two kinds of various concentrations is inputted into micro-fluidic core from above-mentioned two solution inlet port simultaneously with identical flow velocity In piece, mixed with the blood sample in detection cell；

(3) with the red blood cell photo in micro- sem observation and shot detection pond；

(4) from identifying intact red blood cells in each detection cell in red blood cell photo and count.

2, opaque kinetic turbidimetric assay: (a kind of full-automatic determining instrument of osmotic fragility of Yang Jianbo: China, 103033482A [P] .2013-04-10.):

Detecting step is as follows:

(1) be ready to heparin tube, built-in 3.5g/L NaCl-PBS buffer 1.5mL multi-joint cuvette；

(2) instrument samples automatically, and is loaded to the cuvette containing the NaCl-PBS buffer that 1.5mL concentration is 3.5g/L, mixes It is even.

(3) cuvette is pushed into detector by instrument automatically, after detection mixes when first second and last second (empirical value is 40 seconds) The absorbance value of mixed liquor.

(4) hemolysis rate %=(absorbance value when absorbance value-measurement when measuring the first second last second)/measures suction when the first second Shading value × 100%.

3, (Wang Weiwei, Yuan Xiangliang, Fei Qili, Shen Lisong, flow cytometry quickly measure erythrocyte infiltration to flow cytometry Brittle application [J] laboratory medicine, 2016;31 (10), 911-915):

Detecting step is as follows:

It prepares red cell suspension: taking whole blood physiological saline through 2 footworks dilution system；

Above-mentioned suspension is mixed into flow cytometer analysis (gating analytic approach), total number of cells is first obtained, adds certain volume Sterile water (volume different, penetration different), lead to osmotic hemolysis, indicate thin by calculating red blood cell residual percentage The degree of born of the same parents' infiltration haemolysis.Determined by the red blood cell residual percentage range of positive controls, when red blood cell to be measured remains Percentage is lower than judgement osmotic fragility increases in limited time thereon.

4, scattered light urbidmetry (performance rate method): (ShenZhen PuMen Science Co., Ltd, it is a kind of red thin based on scattering turbidimetry technology Born of the same parents' osmotic fragility measuring method: China, CN 105928907A [P] .2016.0907. unauthorized)

Detecting step is as follows:

(1) in the cuvette of scattering turbidimetry analysis module, the buffer salt solution of Hyposmolality is added；

(2) red blood cell is added into above-mentioned cuvette, after stirring and evenly mixing, measures the scattering light value A0 of mixed liquor；

(3) after haemolysis occurs for mixed liquor, before reaching haemolysis equilibrium state, the scattering light value A1 of mixed liquor is measured；

(4) according to formula: hemolysis rate %=(A0-A1)/A0 × 100% calculates hemolysis rate, exports result.

In above-mentioned several method, traditional-handwork method and improvement Sanford method are manual methods, and manual operations is more many and diverse and consumes Duration；And influenced by experimental implementation: it is required that each NaCl solution prepared must be accurate, operating process need to reduce mechanical oscillation, Avoid artificial haemolysis；The osmotic fragility method of inspection based on micro-fluidic chip, instrument cost valuableness (micro-fluidic chip + microscope+computer+software), it is unfavorable for popularizing；It is complicated then to there is instrument cost valuableness, instrumentation in flow cytometry, and needs It wants preliminary experiment to determine suitable penetration, is unsatisfactory for clinical practice use demand.

One tube method greatly reduces operating procedure for manual method, while also shortening detection time, but makes When with the absorbance of 530nm wavelength measurement solution, the red blood cell and cell membrane fragments of non-complete hemolysis can be to the absorbances of solution Interference is generated, causes result inaccurate.

Opaque kinetic turbidimetric assay has done further improvement on the basis of a tube method, has replaced to molten from red blood cell The solvable coloring matter such as free hemoglobin out has the wavelength (530nm) absorbed more by force, using the trip to dissolving out from red blood cell There is the specific wavelength compared with weak absorbing as measurement wavelength (840-1200nm) from the solvable coloring matter such as hemoglobin, test is The impermeable photokinesis turbidity of solution, i.e., opaque kinetic turbidimetric assay (turbidimetry).Although however, the wavelength pair of 840-1200nm The absorption of the soluble substances such as free hemoglobin is weaker, but can still generate absorption, can also impact to testing result.

Scattered light urbidmetry (performance rate method), reflection is a certain (before i.e. non-complete hemolysis) before erythrocyte hemolysis equilibrium state The hemolysis rate at a time point.Quickly due to haemolysis speed of the red blood cell in Hyposmolality buffer salt solution.Different time points Hemolysis rate be it is different, performance rate method to the time control it is more demanding, repeatability is difficult to ensure.In addition, performance rate method can not reflect Haemolysis overall process.

Summary of the invention

To solve the above problems, the application provides a kind of osmotic fragility measuring method and its kit, by this The result that measuring method obtains is more accurate, and can be realized the standardization and consistency of testing result.

According in a first aspect, providing a kind of osmotic fragility measuring method in an embodiment, comprising the following steps:

S1, be separately added into two cuvettes equivalent various concentration Hyposmolality salting liquid, wherein a pipe is incomplete Haemolysis pipe, the Hyposmolality salting liquid being added cannot make whole blood complete hemolysis, and another pipe is complete hemolysis pipe, and what is be added is low Osmotic pressure salting liquid can make whole blood complete hemolysis；

S2, equivalent whole blood is added into two cuvettes respectively, after mixing, detects dissipating for mixed liquor in incomplete haemolysis pipe immediately Penetrate light value A0, in complete hemolysis pipe mixed liquor scattering light value B0；

S3, a certain specific time after haemolysis, measure mixed liquor in incomplete haemolysis pipe scattering light value A1, completely The B1 of mixed liquor in haemolysis pipe；

S4, according to formula: percentage of hemolysis %=(A0-A1)/(B0-B1) × 100% calculates the haemolysis percentage of incomplete haemolysis pipe Rate.

Preferably, the Hyposmolality salting liquid of incomplete haemolysis pipe includes in inorganic salts and buffer and preservative It is at least one.

Preferably, the Hyposmolality salting liquid of complete hemolysis pipe includes at least one in inorganic salts, buffer and preservative Kind.

Preferably, inorganic salts are selected from least one of sodium chloride, potassium chloride, magnesium chloride, sodium sulphate, magnesium sulfate.

Preferably, preservative is selected from least one of Proclin300 and Sodium azide.

Preferably, buffer be selected from phosphate buffer, Glycine-NaOH buffer, Tris- hydrochloride buffer, At least one of HEPES buffer solution.

Preferably, in the Hyposmolality salting liquid of incomplete haemolysis pipe, inorganic salts use concentration for 3.5g/L-4.5g/L's Sodium chloride or other comparable inorganic salts of osmotic pressure.

Preferably, the Hyposmolality salting liquid of complete hemolysis pipe is distilled water or pure water or concentration is 0.1g/L-2g/L Sodium chloride solution or its osmotic pressure and 0.1g/L-2g/L sodium chloride solution other comparable buffer salt solutions of osmotic pressure.

Preferably, the volume ratio of Hyposmolality salting liquid is 1:50-500 in whole blood and incomplete haemolysis pipe.

According to second aspect, a kind of reagent suitable for above-mentioned osmotic fragility measuring method is provided in an embodiment Box, comprising: the Hyposmolality salting liquid of incomplete haemolysis pipe and the Hyposmolality salting liquid of complete hemolysis pipe, incomplete haemolysis pipe Hyposmolality salting liquid include inorganic salts and buffer and at least one of the preservatives, the hyposmosis of complete hemolysis pipe Pressing salting liquid includes inorganic salts, buffer and at least one of the preservatives.

For the turbid skill of transmittance, can thoroughly it disappear according to above-described embodiment due to having used scattering turbidimetry technology Influence except the solvable coloring matter such as the free hemoglobin dissolved out from red blood cell to the absorption of light, keeps result more accurate；By In having used scattering turbidimetry end-point method, detecting scattering light value again after the completion of haemolysis is that testing result is smaller by time effects, It is repeated high；Due to using double-tube method, it is 100% as benchmark using the scattering light changing value of complete hemolysis pipe, spy is calculated Determine the whole blood percentage of hemolysis of the salting liquid pipe of the Hyposmolality of concentration, thus realize the standardization of testing result with it is consistent Property.

Detailed description of the invention

Fig. 1 is haemolysis curve of normal person's whole blood under the sodium chloride solution of various concentration in the embodiment of the present invention one；

Fig. 2 is haemolysis curve of Anemic patients' whole blood under the sodium chloride solution of various concentration in the embodiment of the present invention one.

Specific embodiment

Each step that method describes in specification can also according to those skilled in the art institute can obvious mode into Row sequence is exchanged or adjustment.

Osmotic fragility measuring principle of the invention is as follows:

In a certain specific Hyposmolality salting liquid, whole blood sample is added by a certain percentage, it is red thin in whole blood sample after mixing Born of the same parents are well-dispersed in solution.Since the osmotic pressure in mixed liquor inside red blood cell is higher than the osmotic pressure of Hyposmolality salting liquid, Hydrone is entered inside red blood cell by cell membrane, when reaching a certain level inside hydrone penetrates into red blood cell, red blood cell It expands and ruptures and haemolysis.The concentration of Hyposmolality salting liquid is certain, the ratio of whole blood and Hyposmolality salting liquid is certain In the case where, osmotic fragility is bigger, and percentage of hemolysis is bigger, i.e. osmotic fragility and percentage of hemolysis is at just The relationship of ratio.The scattering value that specific Hyposmolality salting liquid is measured by scattering turbidimetry end-point method, is calculated percentage of hemolysis, from And intuitively reflect osmotic fragility situation.In order to further realize the standardization and consistency of testing result, introduce complete Haemolysis pipe is 100% as benchmark using the scattering light changing value of complete hemolysis pipe, the Hyposmolality for the certain concentration being calculated Salting liquid pipe whole blood percentage of hemolysis it is more accurate.

The present invention provides a kind of osmotic fragility measuring method and its kit.

On the one hand, the present invention provides a kind of osmotic fragility measuring method, comprising the following steps:

S1, be separately added into two cuvettes equivalent various concentration Hyposmolality salting liquid, wherein a pipe is incomplete The Hyposmolality salting liquid of haemolysis pipe, the certain concentration being added cannot make whole blood complete hemolysis, and another pipe is complete hemolysis pipe, Its Hyposmolality salting liquid being added can make whole blood complete hemolysis.Wherein, the Hyposmolality salting liquid packet of incomplete haemolysis pipe Containing inorganic salts and buffer and at least one of the preservatives, i.e., the Hyposmolality salting liquid of incomplete haemolysis pipe may include Inorganic salts and buffer perhaps comprising inorganic salts and preservative or include inorganic salts, buffer and preservative.Inorganic salts choosing From at least one of sodium chloride, potassium chloride, magnesium chloride, sodium sulphate, magnesium sulfate.In one embodiment, inorganic salts use Concentration is the sodium chloride or other comparable inorganic salts of osmotic pressure of 3.5g/L-4.5g/L.Preservative is selected from Proclin300 and folds At least one of nitrogen sodium.Buffer is selected from phosphate buffer, Glycine-NaOH buffer, Tris- hydrochloride buffer At least one of liquid, HEPES buffer solution.

The Hyposmolality salting liquid of complete hemolysis pipe includes inorganic salts, buffer and at least one of the preservatives.Wherein, Inorganic salts are selected from least one of sodium chloride, potassium chloride, magnesium chloride, sodium sulphate, magnesium sulfate.Preservative is selected from Proclin300 At least one of with Sodium azide.It is slow that buffer is selected from phosphate buffer, Glycine-NaOH buffer, Tris- hydrochloric acid At least one of fliud flushing, HEPES buffer solution.In one embodiment, the Hyposmolality salting liquid of complete hemolysis pipe is to steam The sodium chloride of sodium chloride solution or its osmotic pressure and 0.1g/L-2g/L that distilled water or pure water or concentration are 0.1g/L-2g/L is molten Other comparable buffer salt solutions of the osmotic pressure of liquid.

S2, equivalent whole blood, Hyposmolality salting liquid in whole blood and incomplete haemolysis pipe is added into two cuvettes respectively Volume ratio be 1:50-500, the volume ratio of Hyposmolality salting liquid is also 1:50-500 in whole blood and complete hemolysis pipe. After mixing, detect immediately the scattering light value A0 of mixed liquor in incomplete haemolysis pipe, in complete hemolysis pipe mixed liquor scattering light value B0.Wherein, blending manner can be stirring rod stir and evenly mix in mode, magnetic agitation blending manner or bubble blending manner one Kind is a variety of.The light source that wavelength is 400-1200nm can be used in measurement scattering light value, and used light source is laser light source, LED The one of which of light source or common tungsten light source.

S3, a certain specific time after haemolysis, measure mixed liquor in incomplete haemolysis pipe scattering light value A1, The B1 of mixed liquor in complete hemolysis pipe.Above-mentioned specific time according to the difference of the osmotic pressure of Hyposmolality salting liquid and, whole blood Difference, preferred time are 40S or more after sample-adding with the difference of the ratio of the Hyposmolality salting liquid of certain concentration, such as 40S, 50S, 60S, 70S, 80S, 90S, 100S, 120S, 150S, 180S, 200S etc..

S4, it is 100% using the scattering changing value (B0-B1) of complete hemolysis pipe after haemolysis as benchmark, calculates haemolysis hundred Divide rate.According to formula: the haemolysis percentage of incomplete haemolysis pipe is calculated in percentage of hemolysis %=(A0-A1)/(B0-B1) × 100% Rate.

On the other hand, the present invention provides a kind of kit suitable for above-mentioned osmotic fragility measuring method, comprising: The Hyposmolality salting liquid of incomplete haemolysis pipe and the Hyposmolality salting liquid of complete hemolysis pipe, the hyposmosis of incomplete haemolysis pipe Pressing salting liquid includes inorganic salts and buffer and at least one of the preservatives.Wherein, the Hyposmolality of incomplete haemolysis pipe Salting liquid includes that inorganic salts and buffer and at least one of the preservatives, i.e., the Hyposmolality salt of incomplete haemolysis pipe are molten Liquid may include inorganic salts and buffer, perhaps comprising inorganic salts and preservative or include inorganic salts, buffer and preservative. Inorganic salts are selected from least one of sodium chloride, potassium chloride, magnesium chloride, sodium sulphate, magnesium sulfate.In one embodiment, nothing Machine salt uses concentration for the sodium chloride of 3.5g/L-4.5g/L or other comparable inorganic salts of osmotic pressure.Preservative is selected from At least one of Proclin300 and Sodium azide.Buffer be selected from phosphate buffer, Glycine-NaOH buffer, At least one of Tris- hydrochloride buffer, HEPES buffer solution.

Wherein, the Hyposmolality salting liquid of complete hemolysis pipe includes inorganic salts, buffer and at least one of the preservatives. Wherein, inorganic salts are selected from least one of sodium chloride, potassium chloride, magnesium chloride, sodium sulphate, magnesium sulfate.Preservative is selected from At least one of Proclin300 and Sodium azide.Buffer be selected from phosphate buffer, Glycine-NaOH buffer, At least one of Tris- hydrochloride buffer, HEPES buffer solution.In one embodiment, the Hyposmolality of complete hemolysis pipe The sodium chloride solution or its osmotic pressure and 0.1g/L-2g/L that salting liquid is distilled water or pure water or concentration is 0.1g/L-2g/L Sodium chloride solution other comparable buffer salt solutions of osmotic pressure.

Below by specific embodiment combination attached drawing, invention is further described in detail.