Modulus™ Single Tube Fluorometer Method for

Quant-iT™ Protein

1. INTRODUCTION

The Modulus™ Fluorometer from Turner BioSystems in combination with the Quant-iT™ Protein Assay kit from Molecular Probes provides an accurate method for quantitation of protein in small volumes (100 µL). The Quant-iT Protein assay is highly selective for protein and tolerates common contaminants including salts, solvents or DNA, but not detergents.


Figure 1. BSA and Quant-iT Protein analyzed using the Modulus and the Blue Fluorescence Optical Kit. 5 µL of each standard was added to 100 µL of Quant-iT RNA reagent. 100 µL of this mixture was transferred to a minicell cuvette. The Modulus measured raw fluorescence of each dilution.

2. MATERIALS REQUIRED

• Modulus Fluorometer (P/N 9200-000 or 9200-002)
• Blue Fluorescence Optical Kit (P/N 9200-040)
• Quant-iT Protein Assay Kit (Molecular Probes, Q33210)
• Minicell Cuvettes (P/N 7000-950) and Minicell Adaptor (P/N 9200-928)

3. EXPERIMENTAL PROTOCOL

3.1 Reagent Preparation

NOTE: Handling, storage and use of the reagent should be performed in accordance with the product information sheet supplied by Molecular Probes, Inc.

The Quant-iT Protein Reagent is supplied as a 1 mL concentrated dye solution in 1,2-propanediol. On the day of the experiment, equilibrate kit contents to room temperature. Prepare a working solution of the Quant-iT Protein Reagent by making a 1:200 dilution of the concentrated dye solution in Quant-iT Protein buffer. Prepare this solution in a plastic container as the reagent may adsorb to glass surfaces. Protect the working solution from light by covering it with foil or placing it in the dark.

NOTE: For best results, use this solution within 3 hours of its preparation.

3.2 Instrument Set-Up

3.2.1 Power OFF the Modulus. Insert the Blue Fluorescence Optical Kit and Minicell Adaptor according to Operating Manual.

3.2.2 Turn ON the Modulus. Allow a 5-minute warm up period.

3.3 Standard Curve

3.3.1 Add 5 µL of each standard to a microcentrifuge tube containing 100 µL of the Quant-iT Protein Reagent working solution. Mix by inversion.

3.3.2 Transfer 105 µL of each standard to a minicell cuvette.

3.3.3 The default mode for the Modulus is the Raw Fluorescence Mode.
Read all standards in the Raw Fluorescence Mode and record the readings.

3.3.4 Plot amount of the standard vs. fluorescence (FSU) and fit a curve through the data points.

3.4 Sample Analysis

3.4.1 Add 5 µL of each sample to a microcentrifuge tube containing 100 µL of the Quant-iT RNA reagent working solution. Mix by inversion.

3.4.2 Transfer 105 µL of each sample to a minicell cuvette.

3.4.3 Read each sample in the Raw Fluorescence Mode.

3.4.4 Use the standard curve plot from step 3.3.4 to determine the amount of protein in the samples.

4. PATENTS AND TRADEMARKS
Quant-iT is a registered trade--mark of Molecular Probes, Inc.

Modulus is a trademark of Turner BioSystems, Inc.

5. ABOUT MOLECULAR PROBES, INC.
Orders for Molecular Probes' products may be placed by:

Phone: (541) 465-8338 or
Toll-Free: (800) 438-2209 (U.S. and Canada)

Fax: (541) 344-6504 or
Toll-Free: (800) 438-0228 (U.S. and Canada)
E-mail: order@probes.com

Mailing Address:
Molecular Probes, Inc.
PO Box 22010
Eugene, OR 97402-0414 USA

Information on the scientific and technical back-ground of Molecular Probes' products is available from the Technical Assistance Department:

Phone: (541) 465-8353
Fax: (541) 465-4593
E-mail: tech@probes.com
www.probes.com

6. ABOUT TURNER BIOSYSTEMS, INC.

Orders for Turner BioSystems' products may be placed by:

Phone: (408) 636-2400
Toll Free: (888) 636-2401 or
Fax: (408) 737-7919

Contact us via our contact form
www.turnerbiosystems.com

Mailing Address:
Turner BioSystems, Inc.
645 N. Mary Avenue
Sunnyvale, CA 94085

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