CN104330570B - The application of human heat shock protein gp96 in the product of preparation examination hepatopathy - Google Patents
The application of human heat shock protein gp96 in the product of preparation examination hepatopathy Download PDFInfo
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- CN104330570B CN104330570B CN201410535528.9A CN201410535528A CN104330570B CN 104330570 B CN104330570 B CN 104330570B CN 201410535528 A CN201410535528 A CN 201410535528A CN 104330570 B CN104330570 B CN 104330570B
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- 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
- G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
- G01N33/57407—Specifically defined cancers
- G01N33/57438—Specifically defined cancers of liver, pancreas or kidney
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/08—Hepato-biliairy disorders other than hepatitis
- G01N2800/085—Liver diseases, e.g. portal hypertension, fibrosis, cirrhosis, bilirubin
Abstract
The invention discloses the application of human heat shock protein gp96 in the product of preparation examination hepatopathy.The present invention openly detects the application of material in the product preparing examination or the product of auxiliary diagnosis disease or the prognosis of auxiliary judgment disease of human heat shock protein gp96 content.Reference's heat shock protein of the present invention can as a kind of new cirrhosis, liver cancer marker.Kit disclosed by the invention is significant for monitoring after the early screening of cirrhosis, liver cancer, prevention, treatment and treatment.
Description
Technical field
The present invention relates to the application of human heat shock protein gp96 in the product of preparation examination hepatopathy, belong to biomedical sector.
Background technology
Heat shock protein gp96, i.e. GRP94 (glucose-regulatedprotein94) are present in the heat shock protein that eukaryotic cells endoplasmic reticulum middle-molecular-weihydroxyethyl is about 96kDa.It belongs to HSP90 family, is the paralog albumen of kytoplasm HSP90, accounts for 1% of total protein of cell.Under normal physiological conditions, gp96 is positioned in endocytoplasmic reticulum, also exists on the plasma membrane of cell.The signal peptide mediation gp96 molecule of 21 hydrophobic amino acids of gp96N end enters endoplasmic reticulum, under the effect of proteinase, signal peptide is cut in endoplasmic reticulum, kdel receptor on the KDEL sequence that gp96 molecule is held by C and endoplasmic reticulum interacts and makes gp96 molecule be stranded in endoplasmic reticulum, processed in endoplasmic reticulum.Gp96 is high conservative during evolution, and ripe gp96 albumen is made up of four domains: N terminal domains, the region containing ATP-binding domain and conjugated antigen peptide; Charged hinge area, be that gp96 performance is active required, have the change regulating gp96 conformation, charged hinge area can be divided into again the intensive sequence of E/D-and poly-D-lysine repetitive sequence, and the intensive sequence of E/D-contains 1-2 calcium binding sites; Central domain, containing the leucine zipper district and the KEKE district that participate in protein-protein interaction; C terminal domains, containing dimerization tagma, also has the KEDL sequence with receptors bind in endoplasmic reticulum.
Cirrhosis is clinical common chronic liver disease, the diffusivity hepatic lesion that long-term or repeated action is formed by one or more causes of disease.In China, great majority change cirrhosis by hepatitis, and small part is that alcohol and blood fluke cause cirrhosis.Early-phase hepatocirrhosis liver compensation more by force can non-evident sympton, stage with hepatic disorder and portal hypertension for main manifestations, and have multisystem to get involved, often there is the complication such as upper gastrointestinal bleeding, hepatic encephalopathy, scabies secondary infection, hypersplenia, ascites, canceration late period and dead.Therefore, in the urgent need to finding new mark as the Testing index of early-phase hepatocirrhosis.
Primary hepatoma (Hepatocellularcarcinoma, HCC are called for short liver cancer), in worldwide, the incidence of disease comes the 6th, annual global number of the infected is 62.6 ten thousand, and China dies from liver cancer about 110,000 people every year, accounts for 45% of whole world PLC mortality number.Relative to high mortality, the discovery ratio of early liver cancer is but very low.In recent years, although constantly update (comprising excision, transplantation of liver, radiation therapy, interventional therapy, Wicresoft's ablation etc.) for the treatment means of liver cancer, but the general curative effect of liver cancer treatment there is no and significantly improves, recurrence and transfer remain the main factor affecting prognosis in hcc.The detection of current cirrhosis, liver cancer is mainly through iconography, biopsy etc., and SD sensitivity and specificity are not high, and most of patients, once make a definite diagnosis the late period being cirrhosis or liver cancer, loses best treatment period.Therefore, be badly in need of excavating the molecular marker to cirrhosis, liver cancer early screening and simple and easy to do detection method.
Summary of the invention
The object of this invention is to provide the application of human heat shock protein gp96 in the product of preparation examination hepatopathy.
The invention provides the application of material in the product preparing examination or the product of auxiliary diagnosis disease or the prognosis of auxiliary judgment disease detecting human heat shock protein gp96 content;
The amino acid sequence of described human heat shock protein gp96 is as shown in SEQIDNo.2.
In above-mentioned application, the material of described detection human heat shock protein gp96 content comprises the antibody of human heat shock protein gp96 and/or human heat shock protein gp96;
The antibody specific of described human heat shock protein gp96 is the genetic engineering antibody of the monoclonal antibody of human heat shock protein gp96, the polyclonal antibody of human heat shock protein gp96 or human heat shock protein gp96.
In above-mentioned arbitrary described application, the material of described detection human heat shock protein gp96 content also comprises by the reagent in the reagent needed for enzyme-linked immunosorbent assay human heat shock protein gp96 content except the antibody of human heat shock protein gp96 and human heat shock protein gp96 and required instrument.
In above-mentioned arbitrary described application, described enzyme-linked immunosorbent assay is ELISA double antibody sandwich method;
In described ELISA double antibody sandwich method, the antibody specific of human heat shock protein gp96 is that two of the polyclonal antibody of the anti-described human heat shock protein gp96 of the single-chain antibody of human heat shock protein gp96 and/or the polyclonal antibody of human heat shock protein gp96 and/or mark resists;
Two anti-two of the polyclonal antibodies of the anti-described human heat shock protein gp96 of enzyme labeling that are specially of the polyclonal antibody of the anti-described human heat shock protein gp96 of described mark resist;
Two enzymes resisted of the polyclonal antibody of the anti-described human heat shock protein gp96 of described enzyme labeling can develop the color by catalytic substrate;
The single-chain antibody of described human heat shock protein gp96 is specially gp96-scFv-comp antibody, and the sequence of this antibody is as shown in SEQIDNo.1, and the N of this antibody capable specific binding human heat shock protein gp96 holds;
The trade name that the polyclonal antibody of described human heat shock protein gp96 is concrete is goat polyclonal antibodies, and this antibody is purchased from SantaCruzBiotech, and catalog number is SC-1794, and this antibody specific binding is held in the C of human heat shock protein gp96;
Two anti-concrete trade names of the polyclonal antibody of the anti-described human heat shock protein gp96 of described enzyme labeling are the anti-HRP of the anti-goat of rabbit two, and this is two anti-purchased from SantaCruzBiotech, and catalog number is SC-2922.
Recombinant vector containing human heat shock protein gp96 encoding gene, expression cassette, transgenic cell line or recombinant bacterium also belong to protection scope of the present invention as the application of the biomaterial prepared needed for human heat shock protein gp96 in the product preparing examination or the product of auxiliary diagnosis disease or the prognosis of auxiliary judgment disease;
The amino acid sequence of described human heat shock protein gp96 is as shown in SEQIDNo.2.
In above-mentioned application, the encoding gene of described human heat shock protein gp96 is following 1)-3) arbitrary shown in:
1) DNA molecular shown in SEQIDNo.3;
2) under strict conditions with 1) DNA molecule hybridize that limits and the DNA molecular of encoding human heat shock protein gp96;
3) with 1) or 2) DNA molecular that limits has the homogeneity of more than 90% and the DNA molecular of encoding human heat shock protein gp96.
The immunogene of the antibody of the recombinant vector of the encoding gene of the antibody containing human heat shock protein gp96, expression cassette, transgenic cell line or recombinant bacterium or human heat shock protein gp96 or antigen also belong to protection scope of the present invention as the application of the biomaterial needed for the antibody preparing human heat shock protein gp96 in the product of preparation examination or the product of auxiliary diagnosis disease or the prognosis of auxiliary judgment disease;
The antibody of described human heat shock protein gp96 is the genetic engineering antibody of the monoclonal antibody of human heat shock protein gp96 or the polyclonal antibody of human heat shock protein gp96 or human heat shock protein gp96;
The amino acid sequence of described human heat shock protein gp96 is as shown in SEQIDNo.2.
In above-mentioned arbitrary described application, described disease is cirrhosis or liver cancer.
The kit of the prognosis of examination or auxiliary diagnosis disease or auxiliary judgment disease also belongs to a protection scope of the present invention, and this kit comprises the material detecting human heat shock protein gp96 content;
The amino acid sequence of described human heat shock protein gp96 is as shown in SEQIDNo.2.
In mentioned reagent box, the material of described detection human heat shock protein gp96 content comprises the antibody of human heat shock protein gp96 and/or human heat shock protein gp96;
The antibody of described human heat shock protein gp96 is the genetic engineering antibody of the monoclonal antibody of human heat shock protein gp96 or the polyclonal antibody of human heat shock protein gp96 or human heat shock protein gp96.
In above-mentioned arbitrary described kit, described disease is cirrhosis or liver cancer;
Described kit also comprises operation instructions, records as following 1 in instructions) and/or 2) and/or 3) shown in content:
1) with the serum of patient to be measured for detected object, if human heat shock protein gp96 content≤70ng/mL in serum to be checked, this patient candidate to be measured is non-liver cirrhosis patient or non-liver cancer patient; If human heat shock protein gp96 content >=220ng/mL in serum to be checked and <800ng/mL, this patient candidate to be measured is liver cirrhosis patient; If human heat shock protein gp96 content >=500ng/mL in serum to be checked, this patient candidate to be measured is liver cancer patient;
2) with the serum of liver cirrhosis patient for detected object, in serum to be checked, the liver cirrhosis patient of human heat shock protein gp96 content <500ng/mL is judged as the liver cirrhosis patient lower than onset of liver cancer rate in the liver cirrhosis patient 5 years of human heat shock protein gp96 content >=500ng/mL in serum to be checked of onset of liver cancer rate in 5 years in advance;
Or,
In serum, in 5 years of liver cirrhosis patient of human heat shock protein gp96 content <500ng/mL, onset of liver cancer rate is low, and in serum, in 5 years of liver cirrhosis patient of human heat shock protein gp96 content >=500ng/mL, onset of liver cancer rate is high;
Describedly lowly refer to onset of liver cancer rate≤4.3%;
Described height refers to onset of liver cancer rate >=12.7%;
3) with the serum of liver cancer patient for detected object, in serum to be checked, the liver cancer patient of human heat shock protein gp96 content <800ng/mL is judged as that Overall survival is longer than the liver cancer patient of the Overall survival of the liver cancer patient of human heat shock protein gp96 content >=800ng/mL in serum to be checked in advance;
Or,
In serum, the Overall survival of the liver cancer patient of human heat shock protein gp96 content <800ng/mL is 2.8 ± 0.7 years, and in serum, the Overall survival of the liver cancer patient of human heat shock protein gp96 content >=800ng/mL is 1.2 ± 0.4 years;
Described Overall survival is to causing the dead time because of any reason from randomization.
In above-mentioned arbitrary described kit, described enzyme-linked immunosorbent assay is ELISA double antibody sandwich method;
In described ELISA double antibody sandwich method, the antibody specific of human heat shock protein gp96 is that two of the polyclonal antibody of the anti-described human heat shock protein gp96 of the single-chain antibody of human heat shock protein gp96 and/or the polyclonal antibody of human heat shock protein gp96 and/or mark resists;
Two anti-two of the polyclonal antibodies of the anti-described human heat shock protein gp96 of enzyme labeling that are specially of the polyclonal antibody of the anti-described human heat shock protein gp96 of described mark resist;
Two enzymes resisted of the polyclonal antibody of the anti-described human heat shock protein gp96 of described enzyme labeling can develop the color by catalytic substrate;
The single-chain antibody of described human heat shock protein gp96 is specially gp96-scFv-comp antibody, and the sequence of this antibody is as shown in SEQIDNo.1, and the N of this antibody capable specific binding human heat shock protein gp96 holds;
The trade name that the polyclonal antibody of described human heat shock protein gp96 is concrete is goat polyclonal antibodies, and this antibody is purchased from SantaCruzBiotech, and catalog number is SC-1794, and this antibody specific binding is held in the C of human heat shock protein gp96;
Two anti-concrete trade names of the polyclonal antibody of the anti-described human heat shock protein gp96 of described enzyme labeling are the anti-HRP of the anti-goat of rabbit two, and this is two anti-purchased from SantaCruzBiotech, and catalog number is SC-2922.
Reference's heat shock protein gp96 of the present invention can as a kind of new cirrhosis, liver cancer marker.Kit provided by the invention is significant for monitoring after the early screening of cirrhosis, liver cancer, prevention, treatment and treatment.
Accompanying drawing explanation
Fig. 1 is human heat shock protein gp96 content detection typical curve.
Fig. 2 is detection and the statistic analysis result of human heat shock protein gp96 level in healthy population, cirrhosis and liver cancer patient blood serum.
Embodiment
The experimental technique used in following embodiment if no special instructions, is conventional method.
Material used in following embodiment, reagent etc., if no special instructions, all can obtain from commercial channels.
Quantitative experiment in following embodiment, all arranges three repetitions, results averaged.
Coating buffer (being called for short CB) is prepared as follows: by 1.59gNa
2cO
3, 2.93NaHCO
3, 0.2gNaN
3be dissolved in appropriate distilled water, adding distil water is settled to 1L, then adjusts the pH to 9.6 of solution, finally by the membrane filtration of solution with 0.22 μM.
PBS damping fluid is prepared as follows: by 8gNaCl, 0.2gKCl, 3.625gNa
2hPO
412H
2o, 0.24gKH
2pO
4be dissolved in appropriate distilled water, adding distil water is settled to 1L, then adjusts the pH to 7.3 of solution.
PBST damping fluid is prepared as follows: by 8gNaCl, 0.2gKCl, 3.625gNa
2hPO
412H
2o, 0.24gKH
2pO
4be dissolved in appropriate distilled water, add the Tween-20 of 5mL, then adding distil water be settled to 1L, then adjust the pH to 7.3 of solution.
Confining liquid is prepared as follows: 5g bovine serum albumin(BSA) (BSA) is used PBS buffer solution, then is settled to 100mL with PBS damping fluid.
Stop buffer is prepared as follows: joined in distilled water by the 21.7ml concentrated sulphuric acid (98%), and be settled to 200mL with distilled water.
Gp96-scFv-comp antibody according to document " osmanthus obviously; Wu Huiying; Sun Lu; etc. Pichia anomala expression gp96-scFv antibody and biological activity determination [J]. bioengineering journal; 2014; 30 (4): 595 ~ 604. " disclosed method is prepared, this antibody specific binding is held in the N of human heat shock protein gp96.
Goat polyclonal antibodies is purchased from SantaCruzBiotech, and catalog number is SC-1794, and this antibody specific binding is held in the C of human heat shock protein gp96.
The anti-HRP of the anti-goat of rabbit two is purchased from SantaCruzBiotech, and catalog number is SC-2922.
TMB is purchased from eBioscience, and catalog number is 00-4201-56.
Healthy human blood in following embodiment, liver cirrhosis patient blood, liver cancer patient blood or unknown blood sample are provided by Cancer Hospital of Chinese Academy of Medical Sciences and Beijing You An hospital, all blood specimen collections all through the informed consent of blood donors, and all sign Informed Consent Form.
Embodiment 1, employing enzyme linked immunosorbent assay (ELISA) set up human heat shock protein gp96 content detection typical curve
The foundation of human heat shock protein gp96 typical curve adopts ELISA double antibody sandwich method, specific as follows:
One, bag quilt: diluting gp96-scFv-comp antibody (its amino acid sequence is as shown in SEQIDNo.1) to its concentration with coating buffer is 1 μ g/mL, is coated in 96 orifice plates, every hole 100 μ L, is spent the night in 4 DEG C of bags.
Two, wash: use PBST buffer solution 3 times, each 1min.
Three, close: every hole adds 200 μ L confining liquids, is placed in 37 DEG C of closed 2h.
Four, wash: use PBST buffer solution 3 times, each 1min.
Five, application of sample: (its amino acid sequence is as shown in SEQIDNo.2 by human heat shock protein gp96, coding gene sequence is as shown in SEQIDNo.3) dilute with confining liquid, be mixed with the concentration of 20000ng/mL, and carry out 2 times of doubling dilutions with confining liquid, ultimate density is 31.25ng/mL, obtain the gp96 protein solution of serial dilutions, the gp96 protein solution with the serial dilutions of variable concentrations is added 96 orifice plates, every hole 100 μ L, every 1 concentration repeats 3 holes, is experimental port.Blank well (do not add gp96 protein solution and only add 100 μ L confining liquids) is set simultaneously, hatches 2h in 37 DEG C.
Six, wash: use PBST buffer solution 5 times, each 1min.
Seven, detection antibody is added: detect antibody (goat polyclonal antibodies) (confining liquid is 1000:1 with the volume ratio of detection antibody) with confining liquid dilution and obtain detecting antibody-solutions, every hole adds 100 μ L, is placed in 37 DEG C and hatches 1h.
Eight, wash: wash 5 times with PBST, each 1min.
Nine, enzyme labelled antibody is added: obtain two anti-solution with anti-(the anti-HRP of the anti-goat of rabbit two) (volume ratio of confining liquid and the anti-HRP of the anti-goat of rabbit two is 1000:1) of the anti-goat of rabbit two of confining liquid dilution horseradish peroxidase-labeled, every hole adds 100 μ L, is placed in 37 DEG C and hatches 1h.
Ten, wash: wash 7 times with PBST, each 1min.
11, substrate solution colour developing is added: every hole adds 100 μ LTMB substrate solutions, 37 DEG C of lucifuge reaction 10min.
12, cessation reaction: every hole adds 50 μ L stop buffers, room temperature reaction 5min.
13, reading: be placed in by 96 orifice plates on enzyme mark detector, reads the light absorption value in each hole in 450nm place.
14, data analysis: the mean value calculating the light absorption value of 3 repeating holes of each concentration, then deduct the light absorption value of blank well respectively, obtain the OD450 of experimental port.
15, drawing standard curve: with the OD450 of experimental port for horizontal ordinate, the quality of human heat shock protein gp96 is ordinate, and drawing standard curve, obtains typical curve formula.
Human heat shock protein gp96 content detection typical curve as shown in Figure 1.
Fig. 1 shows, when the concentration of human heat shock protein gp96 solution is within the scope of 0 ~ 20000ng/mL, the linear relationship of OD450 and human heat shock protein gp96 quality is good, and related coefficient is 0.9991.Blank well OD450 is below 0.05, and what this method can be special detects that concentration is the human heat shock protein gp96 of 1ng/mL, highly sensitive.
The detection of human heat shock protein gp96 level and statistical study in embodiment 2, healthy population, cirrhosis and liver cancer patient blood serum
One, healthy population, cirrhosis are separated with liver cancer patient blood serum
(1) the blood 5mL of patient is extracted in centrifuge tube according to clinical blood-sampling method, 4 DEG C of standing 3h.
(2) by centrifuge tube in 4 DEG C, the centrifugal 10min of 2000rpm ~ 3000rpm.
(3) pipettor draws the flaxen serum in upper strata, by the serum of collection in-20 DEG C of preservations.
Clinical collection Healthy Human Serum sample 20 example altogether, serum of cirrhosis patients sample 30 example, liver cancer patient blood serum sample 30 example.
Two, the ELISA double antibody sandwich method of embodiment 1 is adopted to detect human heat shock protein gp96 protein content in healthy population, cirrhosis and liver cancer patient blood serum sample.Wherein only the human heat shock protein gp96 solution in step 5 is replaced with serum sample to be checked prepared by step one, obtain the OD450 value of sample, this value is brought into the typical curve formula that embodiment 1 obtains, calculate the quality of the human heat shock protein gp96 of sample, and conversion obtains the content of the human heat shock protein gp96 of sample.
In healthy population, cirrhosis and liver cancer patient blood serum, the detection of human heat shock protein gp96 level and statistic analysis result are as shown in Figure 2.In Fig. 2, the serum of normal representation healthy population; Cirrhosis represents the serum of liver cirrhosis patient; Liver cancer represents the serum of liver cancer patient.
Fig. 2 shows, respectively compared with healthy population, in liver cirrhosis patient, liver cancer patient blood serum, human heat shock protein gp96 content significantly raises and has pole significant difference (P<0.01).
By to a large amount of Healthy Human Serum, serum of cirrhosis patients, liver cancer patient blood serum sample carries out detection and the statistical study of above-mentioned human heat shock protein gp96 level, sets up following judgment criteria:
Human heat shock protein gp96 content≤70ng/mL in the serum of detected object, this detected object candidate is non-liver cirrhosis patient or non-liver cancer patient.
Human heat shock protein gp96 content >=220ng/mL in the serum of detected object <800ng/mL, this detected object candidate is liver cirrhosis patient.
Human heat shock protein gp96 content >=500ng/mL in the serum of detected object, this detected object candidate is liver cancer patient.
The analysis of the accuracy of embodiment 3, judgment criteria
One, unknown blood sample is carried out to the detection of embodiment 2, obtain the detection sample number of Healthy People according to the judgment criteria statistics of embodiment 2, the detection sample number of liver cirrhosis patient, the detection sample number of liver cancer patient.
Two, according to the detection of tumor marker, ultrasonoscopy, CT scan CT, magnetic resonance imaging, radioactivity, the overall target such as liver impedance rheograph or cytolgical examination, unknown blood sample is detected, obtain the actual sample number of Healthy People, the actual sample number of liver cirrhosis patient, the actual sample number of liver cancer patient.
Three, classify overall to testing result by the binomial that goldstandard is determined, as case with contrast (being designated as D+ and D-respectively), the result adopting diagnostic test to detect can be write as positive and negative (being designated as T+ and T-) respectively, and data can arrange into four fold table form, as shown in table 1.Have four possible outcomes in table 1, wherein two is correct, and namely case (cirrhosis or liver cancer patient) is diagnosed as the positive (true positives, TP) and contrasts (Healthy People) and is diagnosed as feminine gender (true negative, TN); Two is wrong, and namely case (cirrhosis or liver cancer patient) is diagnosed as feminine gender (false negative, FN) and contrasts (Healthy People) and is diagnosed as the positive (false positive, FP).
Data 2x2 four fold table diagnosed by table 1.
TP: the actual sample number of liver cirrhosis patient in the detection sample number of liver cirrhosis patient; Or, the actual sample number of liver cancer patient in the detection sample number of liver cancer patient;
FN: the actual sample number of liver cirrhosis patient in the detection sample number of non-liver cirrhosis patient; Or, the actual sample number of liver cancer patient in the detection sample number of non-liver cancer patient;
FP: the actual sample number of liver cirrhosis patient in the detection sample number of Healthy People; Or, the actual sample number of liver cancer patient in the detection sample number of Healthy People;
TN: the actual sample number of Healthy People in the detection sample number of Healthy People.
Four, by the actual sample number of the detection sample number/liver cirrhosis patient of the liver cirrhosis patient specificity (specificity=TN/ (FP+TN)) that to obtain with human heat shock protein gp96 index be criterion examination liver cirrhosis patient, and the sensitivity (sensitivity=TP/ (TP+FN)) that detects is obtained.Result shows, the sensitivity being criterion examination liver cirrhosis patient with human heat shock protein gp96 index and specificity are respectively 89.75% and 100% (P<0.01).
By the actual sample number of the detection sample number/liver cancer patient of the liver cancer patient specificity (specificity=TN/ (FP+TN)) that to obtain with human heat shock protein gp96 index be criterion examination liver cancer patient, and obtain the sensitivity (sensitivity=TP/ (TP+FN)) that detects.Result shows, the sensitivity being criterion examination liver cancer patient with human heat shock protein gp96 index and specificity are respectively 91.37% and 100% (P<0.01).
Therefore, the heat shock protein gp96 contents level in human serum can be used as the index of cirrhosis, liver cancer patient Clinical screening.
The foundation of the human heat shock protein gp96 index in the serum of embodiment 4, prognosis of cirrhotic patients
Found by Spearman correlation analysis, in serum, in human heat shock protein gp96 content and liver cirrhosis patient 5 years there is significant correlation, r=0.67 (P<0.01) in onset of liver cancer rate.Found by t inspection, in serum, human heat shock protein gp96 content is relevant to the Overall survival (OS) of liver cancer patient.In serum human heat shock protein gp96 content <500ng/mL liver cirrhosis patient 5 years in onset of liver cancer rate be 4.3%, in serum human heat shock protein gp96 content >=500ng/mL liver cirrhosis patient 5 years in onset of liver cancer rate be 12.7%, the two significant difference (P<0.01).
Therefore, human heat shock protein gp96 content in serum can be used as the index of cirrhosis prognosis, in serum human heat shock protein gp96 content <500ng/mL liver cirrhosis patient 5 years in onset of liver cancer rate significantly lower than onset of liver cancer rate in the liver cirrhosis patient 5 years of human heat shock protein gp96 content >=500ng/mL in serum.
The foundation of the human heat shock protein gp96 index in the serum of embodiment 5, liver cancer patient prognosis
Found by Spearman correlation analysis, in serum there is significant correlation in the Overall survival (OS) of human heat shock protein gp96 content and liver cancer patient, r=0.56 (P<0.01).Found by t inspection, in serum, human heat shock protein gp96 content is relevant to the Overall survival (OS) of liver cancer patient.In serum, the liver cancer patient OS of human heat shock protein gp96 content <800ng/mL is 2.8 ± 0.7 years (mean+SD), in serum, the liver cancer patient OS of human heat shock protein gp96 content >=800ng/mL is 1.2 ± 0.4 years (mean+SD), the two significant difference (P<0.05).
Therefore, human heat shock protein gp96 content in serum can be used as the index of prognosis in hcc, and in serum, the Overall survival of the liver cancer patient of human heat shock protein gp96 content <800ng/mL is significantly longer than the Overall survival of the liver cancer patient of human heat shock protein gp96 content >=800ng/mL in serum.
Claims (2)
1. for detecting the application of material in the kit of the prognosis of preparation auxiliary judgment disease of human heat shock protein gp96 content;
Described kit comprises the material and operation instructions that detect human heat shock protein gp96 content; Described disease is cirrhosis or liver cancer;
It is characterized in that: as following 1) and/or 2) shown in content:
1) with the serum of liver cirrhosis patient for detected object, in serum to be checked, the liver cirrhosis patient of human heat shock protein gp96 content <500ng/mL is judged as the liver cirrhosis patient lower than onset of liver cancer rate in the liver cirrhosis patient 5 years of human heat shock protein gp96 content >=500ng/mL in serum to be checked of onset of liver cancer rate in 5 years in advance;
Or,
In serum, in 5 years of liver cirrhosis patient of human heat shock protein gp96 content <500ng/mL, onset of liver cancer rate is low, and in serum, in 5 years of liver cirrhosis patient of human heat shock protein gp96 content >=500ng/mL, onset of liver cancer rate is high;
And/or
2) with the serum of liver cancer patient for detected object, in serum to be checked, the liver cancer patient of human heat shock protein gp96 content <800ng/mL is judged as that Overall survival is longer than the liver cancer patient of the Overall survival of the liver cancer patient of human heat shock protein gp96 content >=800ng/mL in serum to be checked in advance;
Or,
In serum, the Overall survival of the liver cancer patient of human heat shock protein gp96 content <800ng/mL is 2.8 ± 0.7 years, and in serum, the Overall survival of the liver cancer patient of human heat shock protein gp96 content >=800ng/mL is 1.2 ± 0.4 years.
2. application according to claim 1, is characterized in that: the material of described detection human heat shock protein gp96 content comprises the antibody of human heat shock protein gp96 and/or human heat shock protein gp96.
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| CN116925219B (en) * | 2023-09-19 | 2023-12-08 | 北京索莱宝科技有限公司 | Antibody of small heat shock protein HSPB1, hybridoma cell strain and application thereof |
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