CN107340394A - The diagnostic system and its operating method of a kind of metastatic carcinoma of bone - Google Patents
The diagnostic system and its operating method of a kind of metastatic carcinoma of bone Download PDFInfo
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- CN107340394A CN107340394A CN201710500874.7A CN201710500874A CN107340394A CN 107340394 A CN107340394 A CN 107340394A CN 201710500874 A CN201710500874 A CN 201710500874A CN 107340394 A CN107340394 A CN 107340394A
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- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
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Abstract
The present invention discloses a kind of diagnostic system of metastatic carcinoma of bone, including bone lancing system, bone tissue section preparation system, immunohistochemical kit and protein detection system；A kind of operating method of above-mentioned diagnostic system is further related to, including：Bone puncture is carried out using medullo-puncture needle under the guiding of CT scanner, to obtain sample；Decalcification processing is carried out using EDTA decalcifying Fluids to the sample of acquisition, carries out the making of bone tissue section；SABC preparation is carried out using immunohistochemical kit, carries out the standardization detection in different primary tumo(u)r sources；Prognosis coherent detection is carried out using protein detection system.The present invention carries out fully optimized in flow, reagent and equipment to the diagnostic system of metastatic carcinoma of bone, extract enough samples, shorten the preparation of bone tissue section, ensure that sample can preferably carry out SABC and genetic test, the standardization detection in different primary tumo(u)r sources and the Protein Detection related to prognosis are realized, improves the accuracy of diagnosis.
Description
Technical field
The invention belongs to biological immunology technical field, more particularly to a kind of diagnostic system of metastatic carcinoma of bone and its operation side
Method.
Background technology
The diagnosis of metastatic carcinoma of bone is divided into clinical diagnosis and pathological diagnosis.Pathological diagnosis is goldstandard, it is necessary to obtain bone tissue,
After the making for carrying out pathological section, Pathomorphology, SABC and the detection of gene mutation, final clear and definite Bone tumour are carried out
Cancer, instruct follow-up treatment.
Clinically the deficiency of pathological diagnosis common technology includes at present：
1) deficiency that bone punctures：It is the first step for obtaining tissue specimen that bone, which punctures, but as invasive operation, is not had at present
Have and carry out extensively in situation of all-level hospitals.The tissue samples amount that bone punctures acquisition is less, and tissue samples amount limits to a certain extent
The accuracy of pathologic finding, it is only capable of carrying out conventional microexamination and the inspection of SABC and gene mutation can not being carried out sometimes
Survey, limit the accuracy of diagnosis, follow-up treatment can not be instructed.
2) deficiency of conventional decalcification method：Decalcification is to make the important step of bone tissue section, to be especially immunized
The bone tissue of histochemical staining.Reagent for decalcification is a lot, and decalcifying agent includes organic acid, inorganic acid, ethylenediamine tetra-acetic acid
(Ethylenediaminetetraacetic acid, EDTA) in addition also has electrolysis decalcification method.Strong organic acid, such as nitric acid
It can be used for the decalcification of close cortex bone with hydrochloric acid, substantial amounts of calcium can be removed in a short time.But these strong acid are to tissue morphology meeting
Harmful effect is produced, decalcification is big to tissue destructive by force, can cause the false negative of immunohistochemical staining and genetic test, can
Mistaken diagnosis and misdiagnosis can be caused.EDTA is a kind of relatively good chelating decalcifying agent, influences minimum to institutional framework, can be preferable
Some enzymes of tissue are preserved, the tissue after EDTA decalcifications can carry out SABC and genetic test.But the method decalcification
Speed is slow, and general take off needs several weeks to the several months.It is used for testing, clinical practice has its limitation.There are improvement EDTA- microwaves at present
The method for radiating joint decalcification, can shorten decalcification time, but heat time bad assurance when making decalcification in this way, and program
Cumbersome, easily excessively decalcification damages to sample.
3) there is no the specific SABC index of bone：The primary tumo(u)r in metastatic carcinoma of bone source is numerous, common pathology result
The diagnosis of metastatic carcinoma can only be confirmed in most cases, be less able to differentiate primary cancer site, can be obtained by SABC inspection
The information of part primary tumor, but combined currently without the SABC of standard, prognosis prediction can not be carried out.
The content of the invention
The defects of it is an object of the invention to overcome in the prior art, there is provided a kind of diagnostic system of metastatic carcinoma of bone and its behaviour
Make method, the links of its diagnostic system to Bone tumour are constantly improved, and can be carried out SABC and genetic test, be carried
The high accuracy of diagnosis, is advantageous to instruct follow-up treatment.
To achieve the above object, the present invention adopts the following technical scheme that：
A kind of bone lancing system, it includes CT scanner and medullo-puncture needle, and the CT scanner is used to guide medullo-puncture needle to carry out
Bone punctures, and the medullo-puncture needle includes needle tubing and the medullo-puncture needle handle affixed with it, extractor and pin are arranged successively in the needle tubing
Core, the center of the medullo-puncture needle handle set cavity for placing the extractor handle and nook closing member handle of superposition, the extraction
Device handle is fixedly connected with one end of the extractor, and the nook closing member handle is fixedly connected with one end of the nook closing member, the pin
Core handle is to be flexibly connected with the medullo-puncture needle handle, wherein, the extractor has convergent C-shaped pawl.
The present invention also provides a kind of diagnostic system of the metastatic carcinoma of bone containing above-mentioned bone lancing system, and it also includes bone tissue
Section preparation system, immunohistochemical kit and protein detection system.
In order to further optimize above-mentioned technical proposal, the technical measures that the present invention is taken also include：
Preferably, the bone tissue section preparation system includes EDTA decalcifying Fluids, and the EDTA decalcifying Fluids are 10%EDTA
Liquid, its component and content are as follows：EDTA 400g, NaOH 170g-174g, distilled water 4L.
Preferably, the immunohistochemical kit includes epithelial origin label and specific epithelial marker thing, it is described on
Skin source property label includes CK, EMA；The specific epithelial marker thing include CEA, CA242, TG, PSA, PSAP, 34 β E12,
AFP、Hepatocyte、β-HCG、CA125。
Preferably, the immunohistochemical kit combines including SABC, and the SABC combination includes：
Prostate cancer：CK、P63、34βE12、PSA、P504S(9AMAC)、AR；
Hyperplasia of prostate/adenoma：CK、P63、34BE12、P504S；
Kidney：CK、EMA、Inhibin、Melan-A、CK7、Vimentin、PAX2、CD10；
The nephroblastoma：WT-1、CK、P53、Ki-67；
Urothelial Carcinoma of Bladder：CK7、CK20、P53、Ki-67、P63；
Seminoma：CK、PLAP、CD117、LCA、OCT3/4；
Stomach cancer：CK7、CK20、Villin、CEA、P53、Ki-67、CDX-2、GST-π、AB/PAS**；
Intestinal cancer：CK7、CK20、Villin、CEA、P53、Ki-67、CDX-2、GST-π、AB/PAS**；
Hepatocellular carcinoma：AFP、CD34、CEA、CK8/18、CK19、Ki-67、P53、HbsAg、Hepatocyte；
Hepatocholangiocarcinoma：CK7、CK20、Villin、AFP、CD34、CEA、CK8/18、CK19、Ki-67、
Hepatocyte、HbsAg；
Cancer of the esophagus：P53、Ki-67、CDX-2、GST-π、P63；
Cancer of pancreas：CEA、P53、Ki-67、CDX-2、GST-π、Villin、CK7、CK20；
Neuroendocrine carcinoma：CD56、CD57、Syn、CgA、CK、CK8/18；
Neuroendocrine lung carcinoma：CD56、CD57、Syn、CgA、CK、CK8/18、CD99、TTF-1、P63、Sp-B；
Thyroid papillary carcinoma：Tg、TTF-1、CK19、HMBE-1、Gelactin-3、Ki-67、TPO；
Medullary carcinoma of thyroid gland：Tg、CT、CD56、CD57、Syn、CgA、CEA；
Infiltrative breast carcinoma：P53、Bcl-2、ER、PR、HER2、E-Cadherin、Ki-67、CK7、GCDFP-15；
Endometrioid tumors/adenocarcinoma of the uterine cervix：Vim、CK、ER、PR、Inhibin、P16、CEA；
Endometrial stroma tumour：Vim、CD10、SMA、Inhibin；
Transitional cell carcinoma:CA125、CK7、CK20、P63；
GCT：CD99、Inhibin、Vim、S100、SMA、CK、CK7、EMA；
Metastatic lung/liver cancer：CK7、CK20、Villin、CEA、TTF-1、CK、AFP、CD34、Hepatocyte；
Metastatic lung/human primary gastrointestinal cancers：CK7、CK20、Villin、CEA、TTF-1、Sp-B、CDX-2；
Metastatic lung/prostate cancer：CK7、CK20、Villin、TTF-1、CK、Sp-B、PSA、PSAP；
Metastatic lung/thyroid cancer：CK7、CK20、Villin、TTF-1、CK、Sp-B、TG；
Shift sexual gland/squamous carcinoma：CKAE1、CKAE3、CK8/18、CK5/6、EMA、CD15、P63；
Metastatic lung/nasopharyngeal carcinoma：CK7、CK20、Villin、TTF-1、CK、Sp-B、EMA、EBV*；
Esophagus poor differentiated carcinoma/neuroendocrine carcinoma：Syn、P63、CgA、CD56、CD99、CKAE3、CK5/6；And
Metastatic ovary mucus/serous carcinoma：CK7、CK20、Villin、CEA、TTF-1、CA125、CDX-2.
Preferably, protein detection system application ImmunohistochemistryMethods Methods detection the differential protein CAPS1 and RPLP2.
On the other hand, the present invention also provides a kind of operating method of the diagnostic system of above-mentioned metastatic carcinoma of bone, and it includes as follows
Step：
Step 1) carries out bone puncture under CT scanner guiding using medullo-puncture needle, to obtain sample；
The sample that step 2) obtains to step 1) carries out decalcification processing using EDTA decalcifying Fluids, will complete decalcification processing
Sample carries out the making of bone tissue section；
Step 3) carries out SABC preparation using the bone tissue section obtained by immunohistochemical kit and step 2), carries out
The standardization detection in different primary tumo(u)r sources；
Step 4) is cut into slices using the bone tissue obtained by protein detection system and step 2) to carry out prognosis coherent detection.
In order to further optimize above-mentioned technical proposal, the technical measures that the present invention is taken also include：
Preferably, the specific steps of sample are obtained in the step 1) to be included：
A) before aspiration biopsy, row x-ray or CT or MRI or PET/CT imageological examinations, the standby diseased region punctured is selected,
Simulate the path punctured；
B) puncture and carried out under CT scanner guiding, the extraction of tissue samples is carried out using medullo-puncture needle；
C) the step b) tissue samples extracted are washed away into clot in heparin saline, chooses slough, will had and represent
The streak or block swollen thing sample of meaning is gently put into soak in formalin and fixed, to prepare sample.
Preferably, the concrete operation step of the step 2) is as follows：The sample that step 1) obtains is put into neutral formal
24h is fixed in woods solution, flowing water rinses 0.5~1 hour, then sample is put into the 10%EDTA decalcifying Fluids of 20 times of volumes, put
In 37 degree of baking ovens, it is placed on shaking table and shakes, it is daily to change EDTA decalcifying Fluids, rinsed 4~6 hours using running water after 3-5 days,
Conventional dehydration, transparent, waxdip, FFPE, serial section number is thick 4 μm, 65 DEG C of roasting pieces 3 hours, will section carry out HE with
IHCS is dyed.
Preferably, the specific steps that in the step 3) prepared by SABC include：
A. conventional section dimethylbenzene dewaxes, graded ethanol aquation；
B. antigen retrieval PH9.0EDTA, boil 20 minutes；Directly washing enters c steps if nonantigenic repair；
C. after natural cooling, fully washing, distilled water flushing three times, are washed three times with PBS, 2 minutes every time；
D. primary antibody is added dropwise, 37 DEG C are incubated 1 hour；
E. washed three times with PBS, 2 minutes every time；
F. secondary antibody, 37 DEG C of incubation half an hour are added dropwise；
G. washed three times with PBS, 2 minutes every time；
H.DAB develops the color 5 minutes, washing；
I. haematoxylin is redyed 2 minutes, washing, the oil blackeite several seconds, washing；
J. gradient alcohol dehydration, dimethylbenzene is transparent, neutral gum mounting.
Compared with prior art, the present invention has advantages below and beneficial effect：
First, bone puncture is carried out under the guiding of CT scanner, can guarantee that and be punctured to privileged site, reduces the secondary work of puncture
With；Puncture needle selects specific medullo-puncture needle, and the specimen amount of acquirement is more, and the positive rate of diagnosis improves, and can meet current genetic test
Needs；
Secondly, for strong acid decalcifying Fluid to big to tissue destructive, EDTA decalcification decalcification speed is slow, it is impossible to meets clinical need
Situation about wanting, We conducted improvement, and bone puncture sample is put into the 10%EDTA decalcifying Fluids of 20 times of volumes, is put in 37 degree of baking ovens
In, be placed on shaking table and shake, it is daily to change EDTA decalcifying Fluids, can be carried out after 3-5 days bone tissue section making this, Ji Nengbao
Card puncturing tissue can preferably carry out SABC and genetic test, shorten decalcification time again, can be widely used in clinic；
Finally, the standard set meal of metastatic carcinoma of bone SABC is established, is standardized according to different primary tumo(u)r sources
Diagnosis；The Protein Detection related to prognosis (CAPS1, RPLP2) is also innovatively added, the two albumen are to apply albumen
The method that group is learned, the differential protein found in metastatic carcinoma of bone and primary bone tumour and normal bone tissues, and pass through U.S. TCGA
Database authentication is related to prognosis.
Brief description of the drawings
Fig. 1 is the structural representation of the diagnostic system of metastatic carcinoma of bone of the present invention；
Fig. 2 is the floor map of medullo-puncture needle of the present invention；
Fig. 3 is the material object of medullo-puncture needle of the present invention and the schematic diagram of sample extraction process；
Reference in figure is：
1st, medullo-puncture needle handle；2nd, needle tubing；3rd, nook closing member handle；4th, nook closing member；5th, extractor handle；6th, extractor.
Embodiment
With reference to embodiment and accompanying drawing, the embodiment of the present invention is further described.Following examples are only
For clearly illustrating technical scheme, and can not be limited the scope of the invention with this.
Embodiment 1
The present embodiment is the medullo-puncture needle in bone lancing system of the present invention.
As shown in Figures 2 and 3, above-mentioned medullo-puncture needle includes needle tubing 2 and the medullo-puncture needle handle 1 affixed with it, in the needle tubing 2
Extractor 6 and nook closing member 4 are arranged successively, and the center upper portion of the medullo-puncture needle handle 1 sets cavity for placing the extraction of superposition
Device handle 5 and nook closing member handle 3, the extractor handle 5 are fixedly connected with one end of the extractor 6, the nook closing member handle 3 with
One end of the nook closing member 4 is fixedly connected, the nook closing member handle 3 and the medullo-puncture needle handle 1 for be flexibly connected (such as snap connection/
It is arranged connection/snap-fastener connection), wherein, the extractor 6 has convergent C-shaped pawl.Above-mentioned medullo-puncture needle handle 1 and nook closing member handle 3
Connection can fix extractor handle 5 positioned between the two.The needle tubing 2 is fixedly connected with medullo-puncture needle handle 1 using connector,
One end of this connector is welded with one end of needle tubing 2, and the other end of this connector uses bolt spiral shell with the bottom of medullo-puncture needle handle 1
Mother's connection.
In this embodiment, not shown in figure, the cavity at the center upper portion of medullo-puncture needle handle can closely be arranged syringe,
Syringe also sets up matched aspiration needle and trochar.
Embodiment 2
The present embodiment is the diagnostic system of metastatic carcinoma of bone of the present invention.
As shown in figure 1, the diagnostic system of above-mentioned metastatic carcinoma of bone includes bone lancing system, bone tissue section preparation system, exempted from
Epidemic disease group kit and protein detection system, worn in the bone lancing system using CT scanner guiding medullo-puncture needle progress bone
Thorn, it includes guiding system (such as puncturing guider and positioner), and its medullo-puncture needle used includes having convergent C-shaped
The extractor of pawl, in favor of the extraction of tissue specimen；The bone tissue section preparation system includes being used for bone tissue section preparation
All reagents and instrument, the reagent includes neutral formalin solution and EDTA decalcifying Fluids etc., and wherein EDTA decalcifying Fluids are
10%EDTA liquid, its component and content are as follows：EDTA 400g, NaOH 170g-174g, distilled water 4L, the instrument include cutting
Blade etc.；The immunohistochemical kit includes being used for all reagents and label and SABC group prepared by SABC
Close, the reagent includes primary antibody, secondary antibody, substrate, developer etc.；The protein detection system application ImmunohistochemistryMethods Methods detection is poor
M-band CAPS1 and RPLP2.
Embodiment 3
The present embodiment is to be used for non-diagnostic and non-treatment purpose using the diagnostic system of the metastatic carcinoma of bone described in embodiment 2
Operating method.
The first step：Bone punctures extraction sample step；
Before aspiration biopsy, row x-ray or CT or MRI or PET/CT imageological examinations, the standby diseased region punctured, mould are selected
Intend the path punctured.Puncture is carried out under the guiding of CT scanner, preoperative row local anaesthesia.
(1) reality tissue sampling in bone：As shown in figure 3, after percutaneous inserting needle, inserting needle is rotated, pin is extracted after penetrating cortex bone
Core, continue to rotate inserting needle, treat that pathological tissues are in the form of a column wedging needle tubing, first horizontal displacement extractor handle, then vertical displacement extraction
Device handle, bottom is firmly pressed into, now lesion columnar structure is extracted the convergent C-shaped pawl constraint of device, 360 ° of rotary handles, column
Pathological tissues will be together removed by detachment, taking-up extractor, pathological tissues, nook closing member is arranged again into the extraction taken out
Device, the needle point of the nook closing member transfer to columnar structure sample backward；
(2) osteolytic lesion is drawn materials in bone：As shown in figure 3, after penetrating cortex of bone, 10ml syringes are taken to extract 3~5ml lifes
Salt solution is managed, trochar different depth different directions suction materials in swollen thing is connect, when trochar is clamped by cortex bone, uses aspiration needle
Entered by the passage of trochar in swollen thing and drawn materials.
The tissue specimen of taking-up should wash away clot in heparin saline, choose slough, will there is the bar rope that represents meaning
Shape or the swollen thing sample of bulk are gently put into soak in formalin and fixed, and should avoid extruding sample.
Medullo-puncture needle of the present invention can be selected according to diseased region and size suitable medullo-puncture needle length (10cm or
15cm) and diameter (9G or 11G), it has the following advantages that：Ensure that 100% success is extracted in the biopsy under any pathologic condition
Rate, minimally-invasive, minimally invasive, substitution open biopsy is easy to be quick, directly extracts tissue, living to bone tumour Multi-layer technology
Aspirate cytology sample, soft tissue bone tissue extract a step and completed after inspection extraction, and sampling diameter ratio is big by 33.3% with model.
Second step：Bone tissue section preparation process；
The sample that the first step is extracted is put into neutral formalin solution and fixes 24h, flowing water rinses 0.5~1 hour, uses
EDTA method decalcifications, running water rinses 4~6 hours after decalcification, conventional dehydration, transparent, waxdip, FFPE, serial section number
, thick 4 μm, 65 DEG C of roasting pieces 3 hours；Section is subjected to HE and IHCS dyeing.
Wherein, the component of EDTA decalcifying Fluids and content are as follows：EDTA 400g, NaOH170g-174g, distilled water 4L, use
Said components are made into 10%EDTA liquid.
3rd step：SABC preparation process；
1. section prepared by second step carries out dimethylbenzene dewaxing, graded ethanol aquation；
2. antigen retrieval PH9.0EDTA, boil 20 minutes；(then directly washing enters the 3rd step for nonantigenic reparation)；
3. after natural cooling, fully washing, distilled water flushing three times, are washed three times with PBS, 2 minutes every time；
4. primary antibody is added dropwise, 37 DEG C are incubated 1 hour；
5. washed three times with PBS, 2 minutes every time；
6. secondary antibody is added dropwise, 37 DEG C of incubation half an hour；
7. washed three times with PBS, 2 minutes every time；
8.DAB develops the color 5 minutes, washing；
9. haematoxylin is redyed 2 minutes, washing, the oil blackeite several seconds, washing；
10. gradient alcohol dehydration, dimethylbenzene is transparent, neutral gum mounting.
Wherein primary antibody and secondary antibody are prepared using this area conventional technical means.
The present invention carries out primary lesion standardizable diagnostic using above-mentioned SABC：Transfer for needing clear and definite source
Knurl, the histiological origin of malignant tumour is aided in determining whether with immunohistochemistry technique.
Common epithelial origin label is as follows：
1st, general label
(1) CK (keratin)
CK hypotypes：CK5/6、CK7、CK8、CK10/13、CK18、CK19、CK20
(2) EMA (epithelial membrane antigen)
2nd, specificity and (or) relative specificity epithelial marker thing
(1) CEA (carcinomebryonic antigen, mark gastrointestinal cancer, adenocarcinoma of lung etc.)
(2) CA242 (tumour correlation Slime antigen, mark cancer of pancreas, knot, carcinoma of the rectum etc.)
(3) TG (thyroglobulin, marking thyroid cancer)
(4) PSA (PSA, being mainly used in the diagnosis of prostate cancer and metastatic prostate cancer)
(5) PSAP (prostate specific acid phosphatase, is mainly used in examining for prostate cancer and metastatic prostate cancer
It is disconnected)
(6) 34 β E12 (high molecular weight cell Keratin, mark prostate basic cell by immunohistochemistry.)
(7) AFP (alpha-fetoprotein, labelling liver cancer, endodermal sinus carcinoma etc.)
(8) Hepatocyte (mark hepatocellular carcinoma)
(9) β-HCG (β human chorionic gonadtropins, mark placenta nutrient leaf cell tumour, germinoma)
(10) CA125 (ovarian cancer antigen)
Conventional SABC combination is as follows：
Uropoiesis and male reproductive system tumour
Prostate cancer：CK、P63、34βE12、PSA、P504S(9AMAC)、AR
Hyperplasia of prostate/adenoma：CK、P63、34BE12、P504S
Kidney：CK、EMA、Inhibin、Melan-A、CK7、Vimentin、PAX2、CD10
The nephroblastoma：WT-1、CK、P53、Ki-67
Urothelial Carcinoma of Bladder：CK7、CK20、P53、Ki-67、P63
Seminoma：CK、PLAP、CD117、LCA、OCT3/4
Digestive system tumor
Stomach cancer：CK7、CK20、Villin、CEA、P53、Ki-67、CDX-2、GST-π、AB/PAS**
Intestinal cancer：CK7、CK20、Villin、CEA、P53、Ki-67、CDX-2、GST-π、AB/PAS**
Hepatocellular carcinoma：AFP、CD34、CEA、CK8/18、CK19、Ki-67、P53、HbsAg、Hepatocyte
Hepatocholangiocarcinoma：CK7、CK20、Villin、AFP、CD34、CEA、CK8/18、CK19、Ki-67、
Hepatocyte、HbsAg
Cancer of the esophagus：P53、Ki-67、CDX-2、GST-π、P63
Cancer of pancreas：CEA、P53、Ki-67、CDX-2、GST-π、Villin、CK7、CK20、
Neuroendocrine tumor
Neuroendocrine carcinoma：CD56、CD57、Syn、CgA、CK、CK8/18
Neuroendocrine lung carcinoma：CD56、CD57、Syn、CgA、CK、CK8/18、CD99、TTF-1、P63、Sp-B
Thyroid papillary carcinoma：Tg、TTF-1、CK19、HMBE-1、Gelactin-3、Ki-67、TPO
Medullary carcinoma of thyroid gland：Tg、CT、CD56、CD57、Syn、CgA、CEA
Mammary gland and female sex organ
Infiltrative breast carcinoma：P53、Bcl-2、ER、PR、HER2、E-Cadherin、Ki-67、CK7、GCDFP-15
Endometrioid tumors/adenocarcinoma of the uterine cervix：Vim、CK、ER、PR、Inhibin、P16、CEA
Endometrial stroma tumour：Vim、CD10、SMA、Inhibin
Transitional cell carcinoma:CA125、CK7、CK20、P63
GCT：CD99、Inhibin、Vim、S100、SMA、CK、CK7、EMA
Metastatic tumo(u)r differentiates
Metastatic lung/liver cancer：CK7、CK20、Villin、CEA、TTF-1、CK、AFP、CD34、Hepatocyte
Metastatic lung/human primary gastrointestinal cancers：CK7、CK20、Villin、CEA、TTF-1、Sp-B、CDX-2
Metastatic lung/prostate cancer：CK7、CK20、Villin、TTF-1、CK、Sp-B、PSA、PSAP
Metastatic lung/thyroid cancer：CK7、CK20、Villin、TTF-1、CK、Sp-B、TG
Shift sexual gland/squamous carcinoma：CKAE1、CKAE3、CK8/18、CK5/6、EMA、CD15、P63
Metastatic lung/nasopharyngeal carcinoma：CK7、CK20、Villin、TTF-1、CK、Sp-B、EMA、EBV*
Esophagus poor differentiated carcinoma/neuroendocrine carcinoma：Syn、P63、CgA、CD56、CD99、CKAE3、CK5/6
Metastatic ovary mucus/serous carcinoma：CK7、CK20、Villin、CEA、TTF-1、CA125、CDX-2.
4th step：Prognosis Protein Detection；
Prognosis Protein Detection process and the 3rd step：SABC in SABC preparation process is identical, and it is applied
ImmunohistochemistryMethods Methods detect the protein for comparing differential expression in Bone tumour tissue and normal bone tissues, have found differential protein
CAPS1, RPLP2, and expressed in U.S. TCGA data base queryings protein expression and the relation of existence, discovery CAPS1 with prognosis just
Correlation, RPLP2 expression and prognosis are negatively correlated.
From above-described embodiment, the present invention is from puncture technique, to the processing of sample, then the selection to SABC, with
And prognosis Protein Detection is set up, it carries out fully optimized in flow, reagent and the equipment to the diagnostic system of metastatic carcinoma of bone, protects
Enough samples can be extracted by having demonstrate,proved, and shorten the preparation time of bone tissue section, ensure that puncturing tissue can be preferably immunized
Groupization and genetic test, the standardization detection in different primary tumo(u)r sources and the Protein Detection related to prognosis are realized, is improved
The accuracy of diagnosis, and be advantageous to instruct follow-up treatment.
The specific embodiment of the present invention is described in detail above, but it is intended only as example, it is of the invention and unlimited
It is formed on particular embodiments described above.To those skilled in the art, it is any to the equivalent modifications that carry out of the present invention and
Substitute also all among scope of the invention.Therefore, the impartial conversion made without departing from the spirit and scope of the invention and
Modification, all should be contained within the scope of the invention.
Claims (10)
1. a kind of bone lancing system, it is characterised in that the bone lancing system includes CT scanner and medullo-puncture needle, the CT scan
Instrument is used to guide medullo-puncture needle to carry out bone puncture, and the medullo-puncture needle includes needle tubing (2) and the medullo-puncture needle handle (1) affixed with it, institute
State and be arranged extractor (6) and nook closing member (4) in needle tubing (2) successively, the center of the medullo-puncture needle handle (1) sets cavity for putting
The extractor handle (5) and nook closing member handle (3) of superposition are put, the extractor handle (5) and one end of the extractor (6) are fixed
Connection, the nook closing member handle (3) are fixedly connected with one end of the nook closing member (4), the nook closing member handle (3) and the medullo-puncture needle hand
Handle (1) is flexible connection, wherein, the extractor (6) has convergent C-shaped pawl.
2. a kind of diagnostic system of the metastatic carcinoma of bone of the bone lancing system containing described in claim 1, it is characterised in that also include
Bone tissue section preparation system, immunohistochemical kit and protein detection system.
3. the diagnostic system of a kind of metastatic carcinoma of bone according to claim 2, it is characterised in that prepared by the bone tissue section
System includes EDTA decalcifying Fluids, and the EDTA decalcifying Fluids are 10%EDTA liquid, and its component and content are as follows：EDTA 400g、NaOH
170g-174g, distilled water 4L.
A kind of 4. diagnostic system of metastatic carcinoma of bone according to claim 2, it is characterised in that the immunohistochemical kit
Including general epithelial origin label and specific epithelial marker thing, the general epithelial origin label include CK,
EMA；The specific epithelial marker thing include CEA, CA242, TG, PSA, PSAP, 34 β E12, AFP, Hepatocyte, β-
HCG、CA125。
A kind of 5. diagnostic system of metastatic carcinoma of bone according to claim 2, it is characterised in that the immunohistochemical kit
Combined including SABC, the SABC combination includes：
Prostate cancer：CK、P63、34βE12、PSA、P504S(9AMAC)、AR；
Hyperplasia of prostate/adenoma：CK、P63、34BE12、P504S；
Kidney：CK、EMA、Inhibin、Melan-A、CK7、Vimentin、PAX2、CD10；
The nephroblastoma：WT-1、CK、P53、Ki-67；
Urothelial Carcinoma of Bladder：CK7、CK20、P53、Ki-67、P63；
Seminoma：CK、PLAP、CD117、LCA、OCT3/4；
Stomach cancer：CK7、CK20、Villin、CEA、P53、Ki-67、CDX-2、GST-π、AB/PAS**；
Intestinal cancer：CK7、CK20、Villin、CEA、P53、Ki-67、CDX-2、GST-π、AB/PAS**；
Hepatocellular carcinoma：AFP、CD34、CEA、CK8/18、CK19、Ki-67、P53、HbsAg、Hepatocyte；
Hepatocholangiocarcinoma：CK7、CK20、Villin、AFP、CD34、CEA、CK8/18、CK19、Ki-67、Hepatocyte、
HbsAg；
Cancer of the esophagus：P53、Ki-67、CDX-2、GST-π、P63；
Cancer of pancreas：CEA、P53、Ki-67、CDX-2、GST-π、Villin、CK7、CK20；
Neuroendocrine carcinoma：CD56、CD57、Syn、CgA、CK、CK8/18；
Neuroendocrine lung carcinoma：CD56、CD57、Syn、CgA、CK、CK8/18、CD99、TTF-1、P63、Sp-B；
Thyroid papillary carcinoma：Tg、TTF-1、CK19、HMBE-1、Gelactin-3、Ki-67、TPO；
Medullary carcinoma of thyroid gland：Tg、CT、CD56、CD57、Syn、CgA、CEA；
Infiltrative breast carcinoma：P53、Bcl-2、ER、PR、HER2、E-Cadherin、Ki-67、CK7、GCDFP-15；
Endometrioid tumors/adenocarcinoma of the uterine cervix：Vim、CK、ER、PR、Inhibin、P16、CEA；
Endometrial stroma tumour：Vim、CD10、SMA、Inhibin；
Transitional cell carcinoma:CA125、CK7、CK20、P63；
GCT：CD99、Inhibin、Vim、S100、SMA、CK、CK7、EMA；
Metastatic lung/liver cancer：CK7、CK20、Villin、CEA、TTF-1、CK、AFP、CD34、Hepatocyte；
Metastatic lung/human primary gastrointestinal cancers：CK7、CK20、Villin、CEA、TTF-1、Sp-B、CDX-2；
Metastatic lung/prostate cancer：CK7、CK20、Villin、TTF-1、CK、Sp-B、PSA、PSAP；
Metastatic lung/thyroid cancer：CK7、CK20、Villin、TTF-1、CK、Sp-B、TG；
Shift sexual gland/squamous carcinoma：CKAE1、CKAE3、CK8/18、CK5/6、EMA、CD15、P63；
Metastatic lung/nasopharyngeal carcinoma：CK7、CK20、Villin、TTF-1、CK、Sp-B、EMA、EBV*；
Esophagus poor differentiated carcinoma/neuroendocrine carcinoma：Syn、P63、CgA、CD56、CD99、CKAE3、CK5/6；And
Metastatic ovary mucus/serous carcinoma：CK7、CK20、Villin、CEA、TTF-1、CA125、CDX-2.
6. the diagnostic system of a kind of metastatic carcinoma of bone according to claim 2, it is characterised in that the protein detection system should
With ImmunohistochemistryMethods Methods detection differential protein CAPS1 and RPLP2.
A kind of 7. operating method of the diagnostic system of the metastatic carcinoma of bone any one of claim 2~6, it is characterised in that
Comprise the following steps：
Step 1) carries out bone puncture under the guiding of CT scanner using medullo-puncture needle, to obtain sample；
The sample that step 2) obtains to step 1) carries out decalcification processing using EDTA decalcifying Fluids, will complete the sample of decalcification processing
Carry out the making of bone tissue section；
Step 3) carries out SABC preparation using the bone tissue section obtained by immunohistochemical kit and step 2), carries out different
The standardization detection in primary tumo(u)r source；
Step 4) is cut into slices using the bone tissue obtained by protein detection system and step 2) to carry out prognosis coherent detection.
8. operating method according to claim 7, it is characterised in that the specific steps bag of sample is obtained in the step 1)
Include：
A) before aspiration biopsy, row x-ray or CT or MRI or PET/CT imageological examinations, the standby diseased region punctured, simulation are selected
The path of puncture；
B) puncture and carried out under CT scanner guiding, the extraction of tissue samples is carried out using medullo-puncture needle；
C) the step b) tissue samples extracted are washed away into clot in heparin saline, chooses slough, will had and represent meaning
Streak or block swollen thing sample be gently put into formalin soak fix, to prepare sample.
9. operating method according to claim 7, it is characterised in that the concrete operation step of the step 2) is as follows：Will
The sample that step 1) obtains, which is put into neutral formalin solution, fixes 24h, and flowing water rinses 0.5~1 hour, then puts sample
Enter the 10%EDTA decalcifying Fluids of 20 times of volumes, be put in 37 degree of baking ovens, be placed on shaking table and shake, it is daily to change EDTA decalcifying Fluids,
Being rinsed 4~6 hours using running water after 3-5 days, conventional dehydration, transparent, waxdip, FFPE, serial section number is thick 4 μm,
65 DEG C of roasting pieces 3 hours, section is subjected to HE and IHCS dyeing.
10. operating method according to claim 7, it is characterised in that SABC is prepared specific in the step 3)
Step includes：
A. conventional section dimethylbenzene dewaxes, graded ethanol aquation；
B. antigen retrieval PH9.0EDTA, boil 20 minutes；Directly washing enters c steps if nonantigenic repair；
C. after natural cooling, fully washing, distilled water flushing three times, are washed three times with PBS, 2 minutes every time；
D. primary antibody is added dropwise, 37 DEG C are incubated 1 hour；
E. washed three times with PBS, 2 minutes every time；
F. secondary antibody, 37 DEG C of incubation half an hour are added dropwise；
G. washed three times with PBS, 2 minutes every time；
H.DAB develops the color 5 minutes, washing；
I. haematoxylin is redyed 2 minutes, washing, the oil blackeite several seconds, washing；
J. gradient alcohol dehydration, dimethylbenzene is transparent, neutral gum mounting.
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