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Goodbye, Bradford Assays!

Drive your research forward with IR-based quantitation.


What if you never had to run another Bradford or BCA? Ever?
With Direct Detect, the first infrared (IR)-based biomolecular quantitation system, just spot 2 L of your sample, blank with buffer, and read! No sample prep, no tedious standard curves, and no messy cuvettes or liquid waste.
The Direct Detect system provides more accurate results without the pitfalls of colorimetric assays. By measuring amide bonds in protein chains, the system accurately determines an intrinsic component of every protein without relying on amino acid composition, dye binding properties or reduction-oxidation (redox) potential. You can evaluate major components of complex mixtures separable from the Amide I & II region, like lipids and carbohydratesmaking lysates and membrane preps easier than ever!

EMD Millipore is a division of Merck KGaA, Darmstadt, Germany

Born from EMD Millipores legacy of innovative protein detection and membrane expertise, this small-footprint system represents a bold new move in quantitation of proteins and other biomolecules with only a tiny sample volume.

How the Direct Detect spectrometer gives you better quantitation:


No need for colorimetric assays, giving you speed, accuracy and reproducibility Works even in the presence of detergents and reducing agents Measures amide bond absorbance, an intrinsic feature of all proteins and peptides Can detect other biomolecules, including nucleic acids, lipids and carbohydrates, for analyzing complex mixtures Requires minimal sample volume (2 L), preserving precious samples

Direct Detect spectrometer works where Bradford and BCA assays fail.
1.6 1.4 Normalized assay signal 1.2 1.0 0.8 0.6 0.4 0.2 0.0
Bradford (1 mg/mL BSA) Micro BCA (20 g/mL BSA) Direct Detect System (1 mg/mL BSA) Thermo Scientic BSA Standard BSA 1x PBS BSA 50 mM DTT BSA 1% SDS

Figure 1. The Direct Detect spectrometer provides accurate and precise results, even in presence of detergent (SDS) and reducing agent (DTT). Using the Direct Detect IR-based quantitation system, calculated concentrations for all the BSA samples matched the prediluted standard (Thermo Scientific BSA Standard). In comparison, the Coomassie Plus (Bradford) Assay provided calculated concentrations that differed greatly in the presence of 1% SDS, and the Micro BCA assay could not provide data in the presence of 50 mM DTT.

Direct Detect spectrometer surpasses current approaches to quantitation.


UV/Vis and Colorimetric Direct Detect Spectrometer Advantages of Direct Detect Spectrometer

Absorbance mode Biomolecules detected Assay compatibility Small sample volume

UV/Vis

Infrared

Intrinsic measurement of amide bonds No extinction coefficient No need for colorimetric/fluorimetric assays Provides confidence of sample purity Peptide analysis (not possible with UV) More information from less sample Saves time More accurate/reproducible Not possible to get accurate results in colorimetric assay with small volume/amount

Protein & Nucleic acid

Protein/Peptide, Nucleic acid, Lipids, Carbohydrates No assay required Yes

Colorimetric (Bradford/BCA) Some instruments

The key to simple analysis: membrane technology


The Direct Detect system exploits a new membrane technology that makes aqueous biological samples compatible with direct infrared analysis. The membrane card employs a hydrophilic polytetrafluoroethylene (PTFE) membrane that is transparent in most of infrared spectral region, so you can spot tiny volumes of your biomolecule solutions directly onto the membrane. No sample prep, no need to run a standard curve every time, no cuvettes, and no liquid waste. Spotted samples are stable in ambient conditions, so youll get reproducible data, whether you read the card in the next hour or next month.

Complex samples? Theyre easy with Direct Detect analysis.


Rat Liver Lysate
0.09 0.00 0.01 0.02 0.03 0.04 0.05 0.06 0.07

Absorbance units

Absorbance units

0.06

0.07

0.08

Lipid

Protein

0.2

Amide I
1,700 1,650 1,600

Amide II
1,550 1,500 1,450

3,100 100

3,000

2,900

2,800

2,700

0.05

Absorbance units

0.05

0.10

0.15

Wavenumber cm-1

Wavenumber cm-1

4,000

3,500

3,000

2,500

2,000

1,500

1,000

500

Wavenumber cm-1

Figure 2. Protein quantitation in presence of lipids within a complex cell lysate is possible because the most intense regions of lipid absorbance are spectrally distinct from the proteins Amide I and Amide II signals.

Requires less sample and more compatible with buffer components compared to colorimetric assays.
Quantitation Approach Assay Range Sample Volume (tube/plate) Amount Protein Required to Measure 1 mg/mL Compatible Incompatible

Direct Detect System

250-5,000 g/mL

2 L

6 g

Chelators Detergents Reducing agents SDS Chelators Most reducing agents Detergents Detergents SDS

Urea Guanidine HCI

Bradford Assay

100-1,500 g/mL

35 L /7 L

21 g

Detergents

BCA Assay Micro BCA Assay Lowry Assay

20-2,000 g/mL 2-40 g/mL 10-1,500 g/mL

50 L /25 L 500 L /150 L 200 L /40 L

30 g 450 g 120 g

Chelators Reducing agents Chelators Reducing agents Chelators Detergents Reducing agents

I loved how intuitive the Direct Detect system was to use and I am especially excited about this technologyI think it will save me a lot of time in the future!
Maya Koike, Ph.D. Postdoctoral researcher and Direct Detect spectrometer user, Childrens Hospital of Philadelphia

Ordering Information
Description Qty Catalogue No.

Direct Detect Spectrometer and Starter Kit Includes Direct Detect Spectrometer Universal Power Adapter Dell Latitude 2120 Netbook and power adapter Direct Detect Software Netbook Stand Spotting Tray Ethernet Cable Direct Detect Assay-free Cards (50/pk) Direct Detect Assay-free Cards (50/pk) Direct Detect Assay-free Cards (50/pk) Direct Detect Assay-free Cards (50/pk)

1 1 1 1 1 1 1 1 1 1 4 8

DDHW00010-WW

DDAC00010-GR DDAC00010-4P DDAC00010-8P

To Place an Order or Receive Technical Assistance In the U.S. and Canada, call toll-free 1-800-645-5476 For other countries across Europe and the world, please visit: www.emdmillipore.com/offices For Technical Service, please visit: www.emdmillipore.com/techservice

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www.emdmillipore.com/directdetect
EMD Millipore and the M mark are trademarks of Merck KGaA, Darmstadt, Germany. Direct Detect is a trademark of Merck KGaA, Darmstadt, Germany. Micro BCA is a trademark of Pierce Biotechnology. All other trademarks are the property of their respective owners. Lit No. PB3664EN00, Rev. A 08/2012 LS SBU-12-05890 Printed in the U.S.A. 2012 EMD Millipore Corporation, Billerica, MA, USA. All Rights Reserved.