DRAQ9™ TECHNICAL DATA SHEET PRODUCT: DRAQ9™ PRODUCT CODES: DR90200; DR91000 PRESENTATION: blue DMSO solution STORAGE: store at -20 °C; use above +20 °C DESCRIPTION DRAQ9™ is a novel far-red fluorescing cell permeant probe that labels membranous and vesicular structures in cytoplasm. It can be combined with common UV-excited and visible-range fluors, including GFP, and is compatible with common cell culture media and buffers. DRAQ9™ enables long-term cell tracking and cell painting for high content phenotypic screening. DRAQ9™ does not label the cell nucleus. SPECTRAL CHARACTERISTICS: Exλmax 605/655 nm Emλmax 698 nm APPLICATIONS • Cell painting / Cell mosaic: for non- a priori screening of cell changes on treatment • Long-term cell tracking – non-toxic, stable labeling of cells over several days • Longitudinal labeling of spheroids – correlation with cellular mass Fig. 1. Spectral profile of DRAQ9™ Fluorescence microscopy & High content screening platforms. BEFORE STARTING Read the SDS. Wear protective clothing, safety goggles and laboratory gloves. Check the concentration of DRAQ9™ stated on the vial label. MATERIALS OFTEN REQUIRED BUT NOT SUPPLIED PBS (azide-free), culture medium (CM), CM without phenol red (“Imaging CM”), paraformaldehyde, warming bath. DETECTING DRAQ9™ SIGNALS (see Fig. 1) DRAQ9™ is optimally excited using yellow/red wavelengths. It is detected with farred filters above 660 nm, using a broad band-pass or long band-pass filter, for example: Chroma Part No. 49019 (Ex. 620/20nm, Em. 665LP, HQ 700/75) Fig. 2. DRAQ9™ labelling of live U2OS cells for 48 h at 2 µM - showing a number of cells in mitosis. PREPARING DRAQ9™ FOR USE DRAQ9™ is supplied at 1 mM in DMSO. As the melting point of DMSO is 19°C it is necessary to warm the product vial in a warming bath to above 19°C before pipetting the required amount of DRAQ9™. Dilute the required amount of DRAQ9™ with PBS or culture medium to a working stock solution of 20 µM (i.e. 1:50), before returning the vial to its outer box and storing in the -20°C freezer. Notes: If procedures demand it, make up diluted (i.e. working conc n.) DRAQ9™ required for up to one day’s lab work e.g. total volume required set up a series of wells. Freeze-thaw cycles do not affect the quality or performance of DRAQ9™, therefore aliquoting is not necessary. Document Ref: DR9.TDS Version #: 004 Issue date: 16/05/23 ©Copyright BioStatus 2017-2023. All Rights Reserved

DRAQ9™ TECHNICAL DATA SHEET EXAMPLE PROTOCOLS The indicator phenol red may introduce unwanted background in live cell imaging of DRAQ9™ and other red/far-red fluorescing reporters and any CM containing it should be replaced with a phenol red-free version of the CM (“Imaging CM”) prior to the start of the period of time-lapse or live-cell endpoint imaging. PROTOCOL 1: LONG-TERM LIVE CELL TRACKING 1. If cells are already in culture medium simply pipette 11% v/v of the 10X working solution of DRAQ9™ into the volume of CM in the well or chamber. Or, preferably.. Add 1 volume of DRAQ9™ working stock...