PolyQ Aggregation Assay to Identify Neuroprotective Effect of a Compound in Worms
Published: April 30, 2023
Abstract
Source: Wang, Q. et al. Caenorhabditis elegans as a Model System for Discovering Bioactive Compounds Against Polyglutamine-Mediated Neurotoxicity. J. Vis. Exp. (2021)
This video describes an in vivo method to screen the protective potential of neuroprotective test compounds in reducing polyQ aggregation in body wall muscle cells of nematodes and ameliorating aggregation-associated proteotoxicity.
Protocol
1. PolyQ aggregation assay Preparation of nematodes for the polyQ aggregation assay Transfer 300-500 synchronized L1 larvae of AM141 to each well of a 48-well plate with 500 µL of S medium containing OP50 (OD570 of 0.7-0.8) and 5 mg/mL of astragalan, typically one well per treatment for one time point. Seal the plate with parafilm and incubate at 20 °C and 120 rpm for 24, 48, 72, and 96 h. Harvest the nematodes in a sterile 1.5 mL microcentrifuge tube and…
Disclosures
The authors have nothing to disclose.
Materials
| C. elegans strain | |||
| AM141 rmIs133 [unc-54p::Q40::YFP] | Caenorhabditis Genetics Center (CGC) | https://cgc.umn.edu/strain/AM141 | |
| E. coli strain | |||
| OP50 | Caenorhabditis Genetics Center (CGC) | https://cgc.umn.edu/strain/OP50 | |
| Reagent | |||
| Sodium azide | Sinopharm Chemical Reagent Co., Ltd. | 80115560 | https://www.reagent.com.cn/goodsDetail/5e981aa807664e26af 551e96ff5f07cd |
| Equipment | |||
| 384-well cell culture plate | Nest Biotechnology Co., Ltd. | 761001 | https://www.cell-nest.com/page94? _l=en&product_id=85 |
| 48-well cell culture plate | Nest Biotechnology Co., Ltd. | 748001 | https://www.cell-nest.com/page94? _l=en&product_id=85 |
| Fluorescence microscope | Guangzhou Micro-shot Optical Technology Co., Ltd. | Mshot MF31-LED | https://www.mshot.com/ article/442.html |
| High-content imaging system | Molecular Devices | ImageXpress | https://www.moleculardevices.com/ products/cellular-imagingsystems#High-Content-Imaging |
| Microcentrifuge | GeneCompany | GENESPEED X1 | https://www.genecompany.com/ index.php/Home/Goods/ goodsdetails/gid/189.html |
| Parafilm M | Sigma-Aldrich | P7793-1EA | https://www.sigmaaldrich.cn/CN/en/product/sigma/p7793?context=product |
| Microscope digital camera | Guangzhou Micro-shot Optical Technology Co., Ltd. | MS60 | https://www.mshot.com/article/677.html |
| Software | |||
| Image acquisition and analysis software | Molecular Devices | MetaXpress | https://www.moleculardevices.com/ products/cellular-imaging-systems/ acquisition-and-analysis-software/ metaxpress |
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Cite This Article
PolyQ Aggregation Assay to Identify Neuroprotective Effect of a Compound in Worms. J. Vis. Exp. (Pending Publication), e21284, doi: (2023).