CN101476999B - Method for fixing organization blocks by methanal stationary liquid - Google Patents
Method for fixing organization blocks by methanal stationary liquid Download PDFInfo
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- CN101476999B CN101476999B CN2009100677800A CN200910067780A CN101476999B CN 101476999 B CN101476999 B CN 101476999B CN 2009100677800 A CN2009100677800 A CN 2009100677800A CN 200910067780 A CN200910067780 A CN 200910067780A CN 101476999 B CN101476999 B CN 101476999B
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Abstract
The invention relates to a method for fixing a tissue block by a formaldehyde fixative. The method comprises the following steps: (1) soaking the tissue block into the neutral or subalkaline formaldehyde fixative in a reaction vessel and stirring the tissue block; (2) using an on-line pH detector to continuously detect the pH value of the fixative, recording the processing time when the pH value of the fixative reaches a subalkaline alarm pH value, and then replacing the fixative; and (3) repeating step (2) until the pH value of the fixative is still more than or equal to 7.0 when the fixative is treated for the time as the last processing time, namely reaching the reaction end point of the tissue block processing. The method is a method for fixing the tissue block by the formaldehyde fixative, which meets the scientific and technological principle and has source basis. The method can be used for commodity type automatic pathologic tissue processing equipment and realize the intelligent and full automatic working mode in the process of fixing the tissue block by the neutral (or subalkaline) formaldehyde fixative .
Description
Technical field
The invention belongs to medical tissue pathological technique professional domain, especially a kind of method of fixing organization blocks by methanal stationary liquid.
Background technology
During the traditional medicine histopathology learns a skill, 4% formaldehyde fixed is liquid-solid decides the Pathologic specimen piece of tissue of drawing materials, generally be to be the formaldehyde fixed liquid immersion treatment piece of tissue repeatedly of 1: 10～20 (g/ml) with w/v, always be no less than 4h with duration, make the fixing sclerosis of piece of tissue, belong to the conventional step-by-step processing method of traditional artificial empirical setting mechanism that follows, the piece of tissue fixation procedure does not still have the monitoring method with tracing basis; In addition, because the difference of each medical institutions' pathological technology room facility condition, personnel specialty technical capability is different with artificial work habit, piece of tissue fixedly treatment process is very not consistent, often cause that fixation of tissue is bad, effect differs, make that morphosis there are differences under the biopsy tissues cell mirror, be the major technique factor that influences the pathological study diagnosis, easily cause the pathological study diagnosis deviation to occur.
Summary of the invention
The objective of the invention is to overcome the deficiencies in the prior art, a kind of method of fixing organization blocks by methanal stationary liquid is provided, make the fixation procedure of piece of tissue meet scientific and technical principle, have tracing basis, the histocyte structure is fixed well stable, can assist the accuracy that improves the pathological study diagnosis.
The present invention solves its technical matters and is achieved through the following technical solutions:
A kind of method of fixing organization blocks by methanal stationary liquid is characterized in that: its method may further comprise the steps:
(1), piece of tissue is soaked in neutrality in the reaction vessel or the alkalescence formaldehyde fixed liquid and stirs;
(2), use online pH to detect the pH value of meter continuous detecting said fixing liquid, when the pH of immobile liquid value reaches slightly acidic warning pH value, write down this time and handle used duration, change immobile liquid then;
(3), repeating step (2), be equal to when once handling used duration until the pH of immobile liquid value and still be 〉=7.0, promptly reach the reaction end that piece of tissue is handled.
And the w/v of described piece of tissue and neutrality or alkalescence formaldehyde fixed liquid is 1: 10～20, g/ml.
And, in the described step (2) volume of each immobile liquid of changing with the first time used immobile liquid volume be that benchmark successively decreases 20% one by one.
And described subalkaline pH value is 7.2～7.6, and described slightly acidic warning pH value is 6.9～6.5.
And the pH value described in the step (3) is 7.0～7.6.
Advantage of the present invention and beneficial effect are:
1. the present invention is in the step-by-step processing process of immobile liquid dosage of repeatedly successively decreasing, when being equal to when once handling used duration, the pH of certain immobile liquid value still is 〉=7.0, the reaction end that tissue protein is fixing is determined in i.e. prompting, makes the piece of tissue fixation procedure have tracing basis.
2. the present invention adopts online pH to detect the pH value of meter continuous detecting immobile liquid in the piece of tissue fixation procedure, and when the pH of immobile liquid value changed the subacidity pH value of setting into by 〉=7.0, immobile liquid was changed in prompting.Therefore the present invention can make the amino acid whose formaldehyde reaction of tissue protein be in neutrallty condition all the time, has guaranteed the chemical property of resultant of reaction, makes the tissue protein stable in properties after fixing.
3. use the inventive method to handle the fixed effect that piece of tissue can improve the histocyte structure, morphosis is well stable under the biopsy tissues cell mirror.Avoided because the phenomenon that the piece of tissue fixed effect that factors such as laboratory facility condition, personnel specialty technical capability cause is poor, morphosis there are differences under the biopsy tissues cell mirror, the auxiliary effectively accuracy that improves the pathological study diagnosis provides great help for the clinician makes correct medical diagnosis on disease and formulates therapeutic scheme.
4. the present invention a kind ofly meets scientific and technical principle, has the method for the fixing organization blocks by methanal stationary liquid of tracing basis, the present invention is used for the automatic pathological tissue treatment facility of commodity-type, can realize the intelligent fully automatic working pattern of neutrality (or alkalescence) fixing organization blocks by methanal stationary liquid process.
Embodiment
The invention will be further described below by specific embodiment, and following examples are descriptive, is not determinate, can not limit protection scope of the present invention with this.
A kind of method of fixing organization blocks by methanal stationary liquid, its step comprises:
(1). be soaked in piece of tissue in neutrality (or alkalescence pH value=7.2～7.6) the formaldehyde fixed liquid in the reaction vessel and stir.Piece of tissue is 1: 10～20 with the w/v of neutral (or alkalescence) formaldehyde fixed liquid, g/ml.Related formaldehyde fixed liquid is the solution of 4% content of formaldehyde that is mixed by commodity formaldehyde reagent or paraformaldehyde and neutrality (or alkalescence) mixed phosphate among the present invention, down with.
(2). use online pH to detect the pH value of meter continuous detecting said fixing liquid, when the pH of immobile liquid value reaches slightly acidic warning pH value (pH value=6.9～6.5), write down this time and handle used duration, change immobile liquid then.
(3). repeating step (2), be equal to when once handling used duration until the pH of immobile liquid value and still be 〉=7.0, i.e. prompting reaches the reaction end that piece of tissue is handled, and piece of tissue is fixing to be finished.
Among the present invention, in the step (2) volume of each immobile liquid of changing with the first time used immobile liquid volume be that benchmark successively decreases 20% one by one.
Embodiment 1
(1). with gross mass is that the piece of tissue of some 15 * 15 * 3mm sizes of 30g is soaked in the 300ml alkalescence formaldehyde fixed liquid in the reaction vessel and stirs.
(2). use online pH to detect the pH value of meter continuous detecting immobile liquid, react a period of time after, when the pH of immobile liquid value reaches slightly acidic warning pH value, write down this time and handle used duration, then immobile liquid is toppled over out.Subacidity warning pH value in the present embodiment is 6.8.
(3). change immobile liquid, place reaction vessel and piece of tissue to proceed reaction and timing the alkalescence formaldehyde fixed liquid of 240ml, handle in the used duration for the first time being not more than,, change immobile liquid and record and handles used duration the second time if the pH value of immobile liquid reaches at 6.8 o'clock.
(4). handle and timing with the alkalescence formaldehyde fixed liquid of 180ml for the third time, handle in the used duration for the second time being not more than, reach at 6.8 o'clock, change immobile liquid and write down and handle used duration for the third time if the pH value of immobile liquid detects.
(5). handle and timing with the alkalescence formaldehyde fixed liquid of 120ml for the 4th time, if be equal to when handling used duration for the third time, it is 7.6 that the pH value of immobile liquid detects, i.e. prompting reaches the reaction end that piece of tissue is handled, the fixing end of piece of tissue.
Embodiment 2
(1). with gross mass is that the piece of tissue of some 15 * 15 * 3mm sizes of 25g is soaked in the 300ml alkalescence formaldehyde fixed liquid in the reaction vessel and stirs.
(2). use online pH to detect the pH value of meter continuous detecting immobile liquid, react a period of time after, when the pH of immobile liquid value reaches slightly acidic warning pH value, write down this time and handle used duration, then immobile liquid is toppled over out.Subacidity warning pH value in the present embodiment is 6.5.
(3). change immobile liquid, place reaction vessel and piece of tissue to proceed reaction and timing the alkalescence formaldehyde fixed liquid of 240ml, handle in the used duration for the first time being not more than,, change immobile liquid and record and handles used duration the second time if the pH value of immobile liquid reaches at 6.5 o'clock.
(4). handle and timing with the alkalescence formaldehyde fixed liquid of 180ml for the third time, handle in the used duration for the second time being not more than, reach at 6.5 o'clock, change immobile liquid and write down and handle used duration for the third time if the pH value of immobile liquid detects.
(5). handle and timing with the alkalescence formaldehyde fixed liquid of 120ml for the 4th time, if be equal to when handling used duration for the third time, it is 7.0 that the pH value of immobile liquid detects, i.e. prompting reaches the reaction end that piece of tissue is handled, the fixing end of piece of tissue.
Embodiment 3
(1). with gross mass is that the piece of tissue of some 15 * 15 * 3mm sizes of 30g is soaked in the 450ml neutral formalin immobile liquid in the reaction vessel and stirs.
(2). use online pH to detect the pH value of meter continuous detecting immobile liquid, react a period of time after, when the pH of immobile liquid value reaches slightly acidic warning pH value, write down this time and handle used duration, then immobile liquid is toppled over out.Subacidity warning pH value in the present embodiment is 6.9.
(3). change immobile liquid, place reaction vessel and piece of tissue to proceed reaction and timing the neutral formalin immobile liquid of 360ml, handle in the used duration for the first time being not more than,, change immobile liquid and record and handles used duration the second time if the pH value of immobile liquid reaches at 6.9 o'clock.
(4). handle and timing with the neutral formalin immobile liquid of 270ml for the third time, handle in the used duration for the second time being not more than, reach at 6.9 o'clock, change immobile liquid and write down and handle used duration for the third time if the pH value of immobile liquid detects.
(5). handle and timing with the neutral formalin immobile liquid of 180ml for the 4th time, if be equal to when handling used duration for the third time, it still is 7.0 that the pH value of immobile liquid detects, i.e. prompting reaches the reaction end that piece of tissue is handled, the fixing end of piece of tissue.
Embodiment 4
(1). with gross mass is that the piece of tissue of some 15 * 15 * 3mm sizes of 30g is soaked in the 600ml neutral formalin immobile liquid in the reaction vessel and stirs.
(2). use online pH to detect the pH value of meter continuous detecting immobile liquid, react a period of time after, when the pH of immobile liquid value reaches slightly acidic warning pH value, write down this time and handle used duration, then immobile liquid is toppled over out.Subacidity warning pH value in the present embodiment is 6.9.
(3). change immobile liquid, place reaction vessel and piece of tissue to proceed reaction and timing the neutral formalin immobile liquid of 480ml, handle in the used duration for the first time being not more than,, change immobile liquid and record and handles used duration the second time if the pH value of immobile liquid reaches at 6.9 o'clock.
(4). handle and timing with the neutral formalin immobile liquid of 360ml for the third time, handle in the used duration for the second time being not more than, reach at 6.9 o'clock, change immobile liquid and write down and handle used duration for the third time if the pH value of immobile liquid detects.
(5). handle and timing with the neutral formalin immobile liquid of 240ml for the 4th time, if be equal to when handling used duration for the third time, it still is 7.0 that the pH value of immobile liquid detects, i.e. prompting reaches the reaction end that piece of tissue is handled, the fixing end of piece of tissue.
Principle of the present invention is:
Research based on biological chemistry and organic chemistry correlation theory is thought, formaldehyde fixed liquid to the fixation mechanism of piece of tissue be formaldehyde can with nearly all building stone generation addition reaction on the tissue protein, especially the alpha-amido in the amino acid generates a kind of metastable dihydroxymethyl compound---N with regard to easy and formaldehyde reaction under normal temperature and neutrality (or alkalescent) condition, N-dihydroxymethyl amino acid, this reaction also claims amino acid whose formaldehyde reaction, is the chemical reaction of representative, main formaldehyde fixed tissue protein.Formaldehyde is as a kind of protein cross agent, and its intermolecular crosslinked action causes the structural change of protein own, makes to organize hardening and follow the string, thereby has suppressed all cell functions, reaches the purpose of fixation of tissue, the hardening of tissue.
R-CH(NH
3 +)-COO
-+2HCHO→R-CH{N(CH
2OH)
2}-COO
-+H
+
Along with proteocrasic chemical reaction carries out, per 2 formaldehyde molecules are in conjunction with falling 1-NH
3 +, just have 1 H
+Be discharged in the immobile liquid, cause H in the immobile liquid
+Concentration increases gradually, and the purpose of use neutral formalin immobile liquid just is to be maintained fixed the pH value stabilization of liquid in the Specialized Theory.But the surge capability of mixed phosphate has certain limit, the H that reaction generates
+After concentration exceeds its surge capability, the pH value of immobile liquid just tends to acidity change, acid solvent condition can not be reacted and be generated the metastable dihydroxymethyl compound of character, therefore in the formaldehyde reaction process of gal4 amino acid, the pH value of monitoring immobile liquid keeps neutrallty condition, can guarantee to generate the chemical property of product, again can be as the good index of changing immobile liquid, can also point out the reaction end of determining that tissue protein is fixing, make the piece of tissue fixation procedure have tracing basis.
Claims (5)
1. the method for a fixing organization blocks by methanal stationary liquid, it is characterized in that: its method may further comprise the steps:
(1), piece of tissue is soaked in neutrality in the reaction vessel or the alkalescence formaldehyde fixed liquid and stirs;
(2), use online pH to detect the pH value of meter continuous detecting said fixing liquid, when the pH of immobile liquid value reaches slightly acidic warning pH value, write down this time and handle used duration, change immobile liquid then;
(3), repeating step (2), be equal to when once handling used duration until the pH of immobile liquid value and still be 〉=7.0, promptly reach the reaction end that piece of tissue is handled.
2. the method for a kind of fixing organization blocks by methanal stationary liquid according to claim 1, it is characterized in that: the w/v of described piece of tissue and neutrality or alkalescence formaldehyde fixed liquid is 1: 10～20, g/ml.
3. the method for a kind of fixing organization blocks by methanal stationary liquid according to claim 1 is characterized in that: in the described step (2) volume of each immobile liquid of changing with the first time used immobile liquid volume be that benchmark successively decreases 20% one by one.
4. the method for a kind of fixing organization blocks by methanal stationary liquid according to claim 1, it is characterized in that: described subalkaline pH value is 7.2～7.6, described slightly acidic warning pH value is 6.9～6.5.
5. the method for a kind of fixing organization blocks by methanal stationary liquid according to claim 1, it is characterized in that: the pH value described in the step (3) is 7.0～7.6.
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| CN2009100677800A CN101476999B (en) | 2009-01-23 | 2009-01-23 | Method for fixing organization blocks by methanal stationary liquid |
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| CN2009100677800A CN101476999B (en) | 2009-01-23 | 2009-01-23 | Method for fixing organization blocks by methanal stationary liquid |
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| CN101476999B true CN101476999B (en) | 2011-03-16 |
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| DE102009025574A1 (en) * | 2009-06-19 | 2010-12-23 | Leica Biosystems Nussloch Gmbh | Method for automatically processing tissue samples in a tissue processor |
Citations (3)
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| RU2004138524A (en) * | 2004-12-28 | 2006-06-10 | Федеральное государственное образовательное учреждение высшего профессионального образовани "Воронежский государственный аграрный университет им. Г.Д. Глинки" (ФГОУ ВПО ВГАУ им. К.Д. Глинки) (RU) | METHOD FOR PREPARING ERYTHROCITAR DIAGNOSTICUM FOR DETERMINING ANTIBODIES TO LIVER TISSUE |
| EP1895287A2 (en) * | 2006-08-29 | 2008-03-05 | Biosepar -Gesellschaft für Medizin- und Labortechnik mbH | Immobiliser for immobilising biological materials |
| CN101324493A (en) * | 2008-07-29 | 2008-12-17 | 孙翔宇 | Method for rapidly manufacturing large tissue specimen slice |
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- 2009-01-23 CN CN2009100677800A patent/CN101476999B/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| RU2004138524A (en) * | 2004-12-28 | 2006-06-10 | Федеральное государственное образовательное учреждение высшего профессионального образовани "Воронежский государственный аграрный университет им. Г.Д. Глинки" (ФГОУ ВПО ВГАУ им. К.Д. Глинки) (RU) | METHOD FOR PREPARING ERYTHROCITAR DIAGNOSTICUM FOR DETERMINING ANTIBODIES TO LIVER TISSUE |
| EP1895287A2 (en) * | 2006-08-29 | 2008-03-05 | Biosepar -Gesellschaft für Medizin- und Labortechnik mbH | Immobiliser for immobilising biological materials |
| CN101324493A (en) * | 2008-07-29 | 2008-12-17 | 孙翔宇 | Method for rapidly manufacturing large tissue specimen slice |
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