Antibody capable of bonding with LINC0266-1
A technology of antibodies and peptides, applied in the field of cancer diagnosis and treatment, can solve the problems of non-existence of antibodies, and achieve the effects of increasing the positive rate, high mortality rate, and short survival period
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Embodiment 1
[0057] Prepare the anti-LINC00266-1 polypeptide antibody, and detect the titer of the prepared anti-LINC00266-1 polypeptide antibody, the specific operation process is as follows:
[0058] (1) The designed antigenic epitope peptide 1 (shown in SEQ ID NO: 2) and antigenic epitope peptide 2 (shown in SEQ ID NO: 3) against the LINC00266-1 polypeptide were synthesized by chemical synthesis (Gill Biochemical (Shanghai) Co., Ltd.), then the synthetic epitope peptide 1 and antigen epitope peptide 2 were coupled with keyhole limpet hemocyanin as immune antigen 1 and immune antigen 2, and were analyzed by high pressure liquid chromatography (HPLC). The immune antigen is separated and purified to make its purity >85%.
[0059] Take 2 healthy New Zealand rabbits with body weight not less than 2kg, named as the rabbit of antigenic epitope peptide 1 and the rabbit of antigenic epitope peptide 2 respectively. 2 to 3 ml of blood from the above-mentioned rabbit ear arteries were collected, a...
Embodiment 2
[0074] Anti-LINC00266-1 antibodies prepared based on epitope peptide 1 and anti-LINC00266-1 antibodies prepared based on epitope peptide 2 were used for specific detection experiments on LINC00266-1 polypeptides. The specific steps are as follows:
[0075] The first step is to construct the LINC00266-1-GFP fusion gene expression vector. First, site-directed mutation of the wild-type GFP (GFPwt) gene translation promoter ATGGTG sequence to ATTGTT (GFPmut). Therefore, the translation of the GFP gene is destroyed, and the translation promoter ATG on the LINC00266-1 gene is used for translation, thereby expressing and producing the LINC00266-1-GFP fusion protein. After transfecting HeLa cells (ATCC, Number: CCL-2) with liposome Lipo2000 (Life technology) for 24 hours, the cells were collected and lysed to prepare protein samples. Using the anti-LINC00266-1 polypeptide antibody prepared based on antigenic epitope peptide 1 and the anti-LINC00266-1 polypeptide antibody prepared bas...
Embodiment 3
[0080] Using the Western blotting method, the anti-LINC00266-1 antibody prepared by the present invention was used to detect the expression level of the LINC00266-1 polypeptide in colon cancer cells with different invasion and metastasis potentials from the same source. The specific process is as follows:
[0081] Two pairs of colon cancer cells SW480 and SW620 (ATCC, Number: CCL-228 and CCL-227, respectively), HTC-116 (ATCC, Number: CCL-247) and HTC-116high (this cell Established by the previous screening in our laboratory), the cells were lysed to prepare protein samples. Using the Western blotting method, the anti-LINC00266-1 polypeptide antibody prepared by the present invention was used to detect the difference in the level of LINC00266-1 polypeptide between the paired cells. The result is as image 3 As shown, the level of LINC00266-1 polypeptide was significantly up-regulated in colon cancer cells SW620 and HTC-116high with high invasion and metastasis potential. In t...
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