Chikungunya ELISA Protocol
酶联免疫吸附法（ELISA）测定基孔肯雅病毒
发布: 2014年02月20日第4卷第4期 DOI: 10.21769/BioProtoc.1049 浏览次数: 10690
评审: Anonymous reviewer(s)
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参见作者原研究论文
The authors used this protocol in:
Jun 2013
Abstract
This protocol will result in the accurate qualitative measurement of anti-Chikungunya virus antibody (Ab) from infected mouse tissue or serum. This assay was developed by Dr. Caitlin Briggs, Arbovax, Inc. Chikungunya is a BL3 agent and should be handled in a biosafety level 3 laboratory under BL3 conditions. This protocol was used in the publication “Chikungunya virus host range E2 transmembrane deletion mutants induce protective immunity against challenge in C57BL/6J mice” (Piper et al., 2013).
Materials and Reagents
- Baby hamster kidney (BHK) cells (ATCC, catalog number: CCL-10 )
- Chikungunya Virus (CHIKV)
- Potassium tartrate
- 15% potassium tartrate in Dulbecco’s phosphate buffered saline (PBS-D) (Sigma-Aldrich)
- 35% potassium tartrate in PBS-D (Sigma-Aldrich)
- Micro BCA protein assay kit (Pierce, catalog number: 23235 )
- Goat anti-mouse IgG HRP (Sigma-Aldrich, catalog number: A5278 )
- ABTS Peroxidase Substrate System (KPL, Kirkegaard & Perry Laboratories, catalog number: 50-62-00 )
- Cavicide disinfectant
- Bleach
- MEM (Life Technologies, catalog number: 11095 )
- Sodium carbonate (Sigma-Aldrich)
- Sodium bicarbonate (Sigma-Aldrich)
- Tween-20 (Fisher Scientific)
- Heat Inactivated Fetal Calf Serum (FCS)
- Sodium Dodecyl Sulfate (SDS) (Sigma-Aldrich)
- Gentamicin sulfate (Life Technologies, catalog number: 15710 )
- Potassium chloride (Sigma-Aldrich)
- Potassium phosphate (monobasic) (Sigma-Aldrich)
- Sodium chloride (Sigma-Aldrich)
- Sodium phosphate (dibasic) (Sigma-Aldrich)
- Completed 1x MEM (see Recipes)
- PBS-D (see Recipes)
- Wash Buffer (see Recipes)
- Blocking Buffer (see Recipes)
- Carbonate Buffer (pH 9.5) (see Recipes)
Equipment
- Laminar flow biosafety cabinet
- 37 °C incubator with 5% CO2
- 96-well plate reader (405 nm absorbance filter)
- 96-well Nunc MaxiSorp plate (Thermo Scientific, catalog number: 44-2404 -2)
- Microscope
- T-75 vented tissue culture flasks (Corning, catalog number: 430641 )
- Serological pipettes
- Pipet-aid
- 50 ml conical tubes
- Test tube rack
- Benchtop centrifuge with removable buckets with gaskets
- Weigh scale with 0.01 g sensitivity
- Ultra Centrifuge
- SW28 rotor and 38 ml and 17 ml buckets
- Beckman Ultra Clear centrifuge tubes (25 x 89 mm) (Beckman Coulter, catalog number: 344058 )
- Seton centrifuge tubes (16 x 102 mm) (Seton Scientific, catalog number: 7076 )
- Small hand held light/flash light
- Parafilm
Procedure
文章信息
版权信息
© 2014 The Authors; exclusive licensee Bio-protocol LLC.
如何引用
Briggs, C. M., Piper, A. and Hernandez, R. (2014). Chikungunya ELISA Protocol. Bio-protocol 4(4): e1049. DOI: 10.21769/BioProtoc.1049.
Download Citation in RIS Format
分类
免疫学 > 抗体分析 > 抗体-抗原相互作用
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