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Immunofluorescence kit for detecting human prostate cancer antigens PSMA and AR-V7 and application
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An AR-V7, prostate cancer technology, applied in biological testing, measuring devices, material testing products, etc., can solve problems such as unclear guiding significance and inability to reflect the expression level of patients
Pending Publication Date: 2022-05-13
CYTTEL BIOSCI BEIJING
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[0005] At present, for castration-resistant prostate cancer, the detection of simple PSMA has no clear guiding significance for the selection of treatment options, and the detection of PSMA and AR-V7 in tissues is faced with the detection of different tissue sections. Due to the heterogeneity of tumor cells, PSMA and AR-V7 on a single film cannot reflect the expression level of PSMA and AR-V7 in the whole patient
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Embodiment 1
[0039] Materials: smears of blood and body fluid samples with negative or positive enrichment, 22RV1 as control cells, LNCaP smears.
[0040] Experimental steps:
[0041] 1. Wash slides with CYP1 for 3 minutes x 3 times, 100-150 μL each time, to ensure that the entire sample area is covered;
[0042] 2. Absorb the excess liquid on the slide, add CYPP for 5 minutes, wash the slide with CYP1 for 3 minutes×1 time; absorb the excess liquid, add 100-150 μl of blocking solution to seal at room temperature for 25-30 minutes;
[0043] 3. Absorb the excess blocking solution, add 100 μl of diluted PSMA antibody, AR-V7 antibody and CD45 antibody, and incubate in a humid box at room temperature in the dark for 1.5 to 2 hours;
[0044] 4. Protect from light, take CYP2 100-150 μL / tablet, wash the film for 3 minutes×3 times, and absorb excess water;
[0045] 5. Add 100 μl of the diluted secondary antibody, and incubate in a wet box in the dark (37°C, 25-30 min); wash CYP2 for 3 min x 4 tim...
Embodiment 2
[0049] Materials: 1 tube of appropriate amount of anticoagulated blood, which is enriched by membrane filtration and then detected for protein
[0052] 2. Add the suspension to the membrane filtration separation tumor cell technology device, and slowly pass through the filter and membrane;
[0053] 3. After the filtration is completed, continue to add 50ml of 0.01M PBS to the membrane filtration device, wash the cell suspension attached around the tube wall into the membrane filtration device, and let it pass through the filter and membrane;
[0054] 4. Fix the cells on the filter membrane;
[0055] 5. Perform the same operation as in Example 1 to detect the protein.
Embodiment 3
[0057] Materials: 1 tube of appropriate amount of anticoagulated blood, which is enriched by microfluidic method and then detected for protein
[0058] Experimental steps:
[0059] The appropriate amount of blood drawn was enriched by microfluidic chips of various principles, and the enriched samples were detected by protein immunofluorescence.
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Abstract
The invention relates to the technical field of cancerbiology and liquid biopsy, in particular to a human prostatecancerantigen PSMA and androgenreceptor splicing mutant AR-V7 immunofluorescence detection kit which is suitable for detection of different types of samples PSMA and AR-V7, and further relates to application of the kit in detection of PSMA and AR-V7. The kit can be used for detecting the expression levels of PSMA and AR-V7 in prostatecancer non-humoral rare karyocytes so as to provide instructive opinions for medication of prostate cancer patients. Non-humoral rare karyocytes of a patient are typed through specific antibodies aiming at PSMA and AR-V7, and reference opinions are provided for medication of prostatic cancer through the number of PSMA and AR-V7 positive cells and the expression level and intracellular localization of AR-V7 of the PSMA and AR-V7 positive cells. The detection of the immunofluorescence specimen can be suitable for non-humoral rare karyocyte samples obtained by different methods through enrichment, including but not limited to a negative enrichment method, a positive enrichment method, a density gradientseparation method, a membrane filtration method, a microfluidic method and the like.
Description
technical field [0001] The invention relates to the technical fields of cancer biology and liquid biopsy, in particular to an immunofluorescence kit and application for detecting human prostate cancer antigens PSMA and AR-V7. Background technique [0002] PSMA (Prostate-specific membrane antigen, PSMA), the prostate-specific membrane antigen, is a type II transmembrane glycoprotein composed of 750 amino acids, mainly expressed in prostate epithelial cells, and has high prostate cancer specificity. Relevant studies have pointed out that PSMA can be detected in 94.1% of prostate cancertissue sections, and its expression is closely related to tumor grade. Further studies have shown that the use of PSMA radioligands can quickly and accurately detect early disease occurrence, recurrence, and metastasis. [0003] AR-Vs (androgenreceptor splice variants, AR-Vs) refers to the splice mutants produced during the abnormal splicing or gene recombination of androgenreceptor mRNA, whi...
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