BSI PD ISO/TS 19006:2016
$142.49
Nanotechnologies. 5-(and 6)-Chloromethyl-2’,7’ Dichlorodihydrofluorescein diacetate (CM-H2DCF- DA) assay for evaluating nanoparticle-induced intracellular reactive oxygen species (ROS) production in RAW 264.7 macrophage cell line
| Published By | Publication Date | Number of Pages |
| BSI | 2016 | 32 |
This document describes how to test and evaluate results obtained from in vitro ROS generation in RAW 264.7 macrophage cells exposed to nano-objects, nanoparticles, their aggregates and agglomerates using the CM-H2DCFDA assay.
The protocol in this document is limited to use of a 24 well plate so if other plates were to be used, volumes would need to be adjusted and the protocol steps validated to ensure confidence in the test results.
PDF Catalog
| PDF Pages | PDF Title |
|---|---|
| 6 | Foreword |
| 7 | Introduction |
| 9 | 1 Scope 2 Normative references 3 Terms and definitions |
| 11 | 4 Symbols and abbreviated terms 5 Materials |
| 13 | 6 Technical equipment 7 Nanoparticle sample preparation |
| 14 | 8 Preparations 8.1 General |
| 15 | 8.2 Flow cytometry calibration 8.3 Experimental culture medium 8.4 Reagent preparation 8.5 Preparation of cell stock culture 8.6 Preparing culture for experiments |
| 16 | 8.7 Verification of healthy cell growth |
| 17 | 8.8 Evaluation of nanoparticle interference 8.9 Control preparation 8.9.1 General 8.9.2 Control description |
| 18 | 8.9.3 Sin-1 stock solution preparation (1 mM) 9 Evaluation of nanoparticle impact on ROS generation in cells 9.1 Prepare cells in the 24 well plates 9.2 Dose the cells with nanoparticles and controls |
| 19 | 9.3 Expose the cells to CM-H2DCF-DA Assay |
| 20 | 9.4 Incubate the cells with CM-H2DCF-DA 9.5 Flow cytometry analysis 10 Data analysis and results |
| 22 | Annex A (informative) Alternate cell lines |
| 23 | Annex B (informative) Alternate fluorescence characterization techniques |
| 24 | Annex C (informative) Suspension preparation and characterization |
| 25 | Annex D (informative) Example experimental data from RAW 264.7 |
| 28 | Bibliography |
