Nov 01, 2023
- alexron 1
- 1University of Cambridge
Protocol Citation: alexron 2023. MTT assay. protocols.io https://dx.doi.org/10.17504/protocols.io.e6nvwdoqwlmk/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: October 31, 2023
Last Modified: November 01, 2023
Protocol Integer ID: 90238
Keywords: protocol for mtt cytotoxicity, mtt cytotoxicity, sy5y cell, mtt, assay
Abstract
Protocol for MTT cytotoxicity assay on SH-SY5Y cells.
Materials
- Medium: DMEM/F-12, GlutaMAXTM (ThermoFisher Scientific) + 10% Fetal Bovine Serum (GibcoTM)
- Trypsin-EDTA (0.25%) (ThermoFisher Scientific)
- PBS, pH 7.4 (ThermoFisher Scientific)
- RPMI medium (ThermoFisher Scientific)
- MTT kit (Abcam)
- 96-well plates, PS, F-bottom (Greiner Bio-One)
- Plate shaker
- Plate reader for absorbance measurement
General
Culture human SH-SY5Y cells at 37 °C and 5% CO2
Day 1
Detach 80–90% confluent cells with Trypsin-EDTA
Count cells and seed 10000 cells/well
Incubate at 37 °C and 5% CO2 for 24 h
Day 2
Aspirate medium from cells
Add in protein solutions
Place back at 37 °C and 5% CO2 for 24 h
Day 3
Dilute MTT 1:10 in RPMI medium
Aspirate medium from cells
Add 100 µL MTT/RPMI solution per well
Place back at 37 °C and 5% CO2 for 4 h
Aspirate medium from cells
Add 100 µL stop solution per well
Incubate at 37 °C, 500 rpm on plate shaker
Read out absorbance at 570 nm on plate reader