Carboxyl-terminal specific anti-human amyloid protein monoclonal antibody gene and its encoded polypeptide and application
A monoclonal antibody and amyloid technology, which is applied in the field of genetic engineering, can solve the problems of unsatisfactory differential diagnosis and treatment target effect, no Aβ42 monoclonal antibody, and inability to specifically recognize Aβ2 oligomers, etc., and achieve immune effect. Good, high titer of immune serum, and the effect of reducing side effects
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0033] Example 1 Preparation of carboxy-terminal specific anti-human amyloid monoclonal antibody
[0034] 1. Test materials
[0035] 1. Aβ(35-42)(MVGGVVIA) (synthesized by Shanghai Sangon Bioengineering Co., Ltd.)
[0036] 2. Aβ42 peptide (DAEFRHDSGYEVHHQKLVFFAEDVGSNKGAIIGLMVGGVVIA) and Aβ40 peptide (DAEFRHDSGYEVHHQKLVFFAEDVGSNKGAIIGLMVGGVV) (synthesized by Shanghai Sangon Bioengineering Co., Ltd.)
[0037] 3. Hexafluoroisopropanol (1,1,1,3,3,3-hexafluoro-2-propannol, HFIP) (purchased from Sigma)
[0038] 4. Anhydrous dimethyl sulfoxide (DMSO) (purchased from Sigma)
[0039] 5. F12 medium (purchased from Sigma Company)
[0040] 6. Complete Freund's adjuvant (purchased from Sigma)
[0041] 7. Incomplete Freund's adjuvant (purchased from Sigma)
[0042] 8. RPMI1640 culture medium (purchased from Sigma Company)
[0043] 9. PEG4000 (polyethylene glycol) (purchased from Sigma)
[0044] 10. Fetal bovine serum (provided by Hangzhou Sijiqing Bioengineering Materials Co., Ltd.) ...
Embodiment 2
[0076] Embodiment 2 Indirect ELISA experiment
[0077] 1. Test materials
[0078] 1. ELISA plate (manufactured by SUNRISE)
[0079] 2. HRP-labeled goat anti-mouse IgG (purchased from Beijing Zhongshan Jinqiao Biotechnology Co., Ltd.)
[0080] 2. Test method
[0081] 1. Coating: take 10 μg / ml Aβ40 and Aβ42 oligomeric mixture as the coating antigen, dissolve in the coating buffer (pH9.6, 0.05mol / L carbonate buffer), add to the microtiter plate, each Well 100 μl, overnight at 4°C.
[0082] 2. PBS-T (Weighing NaCl8g, KCl0.2g, NaCl 2 HPO 4 1.44g, KH 2 PO 4 0.44g, Tween-200.05ml, add ddH 2 0 to 1 L, adjust the pH to 7.2-7.4) Wash the plate: 3 times, 5 min each time.
[0083] 3. Blocking: add PBS-T containing 0.2% BSA, 100 μl per well. Treat at 37°C for 2h.
[0084] 4. Add the carboxy-terminal specific anti-human amyloid monoclonal antibody diluted in PBS-T times to a 96-well plate, 100 μl per well. Treat at 37°C for 2h. At the same time, a negative control and a positiv...
Embodiment 3
[0098] Example 3 Western blot (Western blot) experiment
[0099] 1. Test method
[0100] Preparation of detection antigen Aβ oligomerization mixture: as shown in Example 1.
[0101] Take 5-10 μg sample, 5×sample buffer, mix well and load the sample, first make the protein pass through the stacking gel with a voltage of 100V. When the sample enters the separating gel, adjust the voltage to keep it constant at 120V. When the bromophenol blue swims to the bottom of the gel, end the electrophoresis, remove the gel, and stain it with Coomassie Brilliant Blue R-250 routinely; put the gel and nitrocellulose membrane into containers containing blotting buffer Equilibrate in the chamber for 10 minutes, put filter paper, gel, NC membrane, and filter paper in order to form a "sandwich" shape, pour the transfer buffer, with the gel side facing the negative electrode and the NC membrane facing the positive electrode, carefully avoiding and driving away air bubbles. Turn on the power, ma...
PUM
Abstract
Description
Claims
Application Information
- R&D Engineer
- R&D Manager
- IP Professional
- Industry Leading Data Capabilities
- Powerful AI technology
- Patent DNA Extraction
Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.
© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com