Thermo NanoDrop One User Manual

NanoDrop Micro-UV/Vis Spectrophotometers

NanoDrop One

User Guide

269-309101 Revision B July 2016

©2015- 2016 Thermo Fisher Scientific Inc. All rights reserved.

DYMO and LabelWriter are either trademarks or registered trademarks of Newell Rubbermaid in the United
States and/or other countries. Wi-Fi is either a trademark or a registered trademark of Wi-Fi Alliance in the
United States and/or other countries. Bluetooth is either a trademark or a registered trademark of Bluetooth
Special Interest Group. Windows is either a trademark or a registered trademark of Microsoft Corporation in
the United States and/or other countries. All other trademarks are the property of Thermo Fisher Scientific inc.
and its subsidiaries.

For U.S. Technical Support, please contact:

Unity Lab Services
Part of Thermo Fisher Scientific
5225 Verona Road

For International Support, please contact:

Thermo Fisher Scientific
Telephone: [phone]
E-mail: [e-mail]

Madison WI 53711-4495 U.S.A.
Telephone: [phone]
E-mail: [e-mail]

Thermo Fisher Scientific Inc. provides this document to its customers with a product purchase to use in the
product operation. This document is copyright protected and any reproduction of the whole or any part of this
document is strictly prohibited, except with the written authorization of Thermo Fisher Scientific Inc.

The contents of this document are subject to change without notice. All technical information in this
document is for reference purposes only. System configurations and specifications in this document supersede
all previous information received by the purchaser.

This document is not part of any sales contract between Thermo Fisher Scientific Inc. and a purchaser. This
document shall in no way govern or modify any Terms and Conditions of Sale, which Terms and Conditions of
Sale shall govern all conflicting information between the two documents.

For Research Use Only. This instrument or accessory is not a medical device and is not intended to be used
for the prevention, diagnosis, treatment or cure of disease.

WARNING Avoid an explosion or fire hazard. This instrument or accessory is not
designed for use in an explosive atmosphere.

C

Contents

Chapter 1 About the NanoDrop One Spectrophotometer. . . . . . . . . . . . . . . . . . . . . . . . . . . . . .1

Instrument Models and Features . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2

Optional Accessories . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5

Register Your Instrument . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

Update Software . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

Chapter 2 Applications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .9

Detection Limits for All Applications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

Measure dsDNA, ssDNA or RNA. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13

Measure dsDNA, ssDNA or RNA. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13

Nucleic Acid Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16

Setting for Nucleic Acid Measurements. . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17

Calculations for Nucleic Acid Measurements. . . . . . . . . . . . . . . . . . . . . . . . . 18

Measure Microarray. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

Measure Microarray Samples. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

Microarray Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27

Settings for Microarray Measurements. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28

Calculations for Microarray Measurements . . . . . . . . . . . . . . . . . . . . . . . . . . 32

Measure using a Custom Factor. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35

Measure Nucleic Acid using a Custom Factor . . . . . . . . . . . . . . . . . . . . . . . . 35

Custom Factor Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 37

Settings for Nucleic Acid Measurements using a Custom Factor . . . . . . . . . . 39

Detection Limits for Nucleic Acid Measurements using a Custom

Factor. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39

Measure Oligo DNA or Oligo RNA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41

Measure Oligo DNA or Oligo RNA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 41

Oligo Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45

Settings for Oligo DNA and Oligo RNA Measurements . . . . . . . . . . . . . . . . 47

Detection Limits for Oligo DNA and Oligo RNA Measurements . . . . . . . . . 48

Calculations for Oligo DNA and Oligo RNA Measurements . . . . . . . . . . . . 49

Thermo Scientific NanoDrop One User Guide iii

Contents

Measure Protein A280. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53

Measure Protein Concentration at A280 . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53

Protein A280 Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 57

Settings for Protein A280 Measurements. . . . . . . . . . . . . . . . . . . . . . . . . . . . 59

Detection Limits for Protein A280 Measurements. . . . . . . . . . . . . . . . . . . . . 64

Calculations for Protein A280 Measurements . . . . . . . . . . . . . . . . . . . . . . . . 65

Measure Proteins and Labels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69

Measure Labeled Protein Samples . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69

Proteins & Labels Reported Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72

Settings for Proteins and Labels Measurements . . . . . . . . . . . . . . . . . . . . . . . 74

Detection Limits for Proteins and Labels Measurements . . . . . . . . . . . . . . . . 76

Calculations for Proteins and Labels Measurements. . . . . . . . . . . . . . . . . . . . 77

Measure Protein A205. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79

Measure Protein Concentration at A205 . . . . . . . . . . . . . . . . . . . . . . . . . . . . 79

Protein A205 Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 82

Settings for Protein A205 Measurements. . . . . . . . . . . . . . . . . . . . . . . . . . . . 83

Calculations for Protein A205 Measurements . . . . . . . . . . . . . . . . . . . . . . . . 85

Measure Protein BCA . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 87

Measure Total Protein Concentration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 87

Protein BCA Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 96

Settings for Protein BCA Measurements . . . . . . . . . . . . . . . . . . . . . . . . . . . 100

Measure Protein Bradford . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 101

Measure Total Protein Concentration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 101

Protein Bradford Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 106

Settings for Protein Bradford Measurements . . . . . . . . . . . . . . . . . . . . . . . . 109

Measure Protein Lowry . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111

Measure Total Protein Concentration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 111

Protein Lowry Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 114

Settings for Protein Lowry Measurements . . . . . . . . . . . . . . . . . . . . . . . . . . 118

Measure Protein Pierce 660. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 119

Measure Total Protein Concentration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 119

Protein Pierce 660 Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 124

Settings for Protein Pierce 660 Measurements . . . . . . . . . . . . . . . . . . . . . . . 127

Measure OD600 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 129

Measure OD600 . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 129

OD600 Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 133

Settings for OD600 Measurements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 134

Calculations for OD600 Measurements . . . . . . . . . . . . . . . . . . . . . . . . . . . 137

Measure Custom . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139

Measure using a Custom Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139

Delete Custom Method. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 143

Custom Method Reported Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 144

iv NanoDrop One User Guide Thermo Scientific

Contents

Measure UV-Vis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147

Measure UV-Vis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 147

UV-Vis Reported Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 150

Settings for UV-Vis Measurements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 152

Measure Kinetics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 155

Measure Kinetics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 155

Create Kinetics Method. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 158

Edit Kinetics Method . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 159

Kinetics Reported Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 161

Settings for Kinetic Measurements. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 166

Chapter 3 Learning Center . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .175

Micro-Volume Sampling—How it Works. . . . . . . . . . . . . . . . . . . . . . . . . . . . 176

Set Up the Instrument. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 178

Measure a Micro-Volume Sample . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193

Measure a Sample Using a Cuvette . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 200

Prepare Samples and Blanks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 204

Basic Instrument Operations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 210

NanoDrop One Home Screen. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 211

NanoDrop One Measurement Screens . . . . . . . . . . . . . . . . . . . . . . . . . . . . 215

NanoDrop One Data Viewer . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 222

NanoDrop One General Operations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 229

Instrument Settings . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 236

Acclaro Sample Intelligence. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 240

NanoDrop One Viewer Software . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 248

Viewer Home Screen. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 249

Manage Experiments and Associated Data. . . . . . . . . . . . . . . . . . . . . . . . . . 251

Manage Identifiers on a PC . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 260

Manage Custom Methods . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 265

Multimedia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 277

Chapter 4 Maintaining Your Instrument . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .279

Maintenance Schedule. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 280

Cleaning the Touchscreen . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 281

Maintaining the Pedestals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 282

Cleaning the Pedestals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 282

Reconditioning the Pedestals . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 285

Decontaminating the Instrument . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 287

Maintaining the Cuvette Sampling System . . . . . . . . . . . . . . . . . . . . . . . . . . . 290

Instrument Diagnostics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 291

Intensity Check . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 291

Performance Verification. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 293

Pedestal Image Check . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 298

Thermo Scientific NanoDrop One User Guide v

Contents

Chapter 5 Safety and Operating Precautions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .301

Operating Precautions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 302

Safety Information. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 303

Chapter 6 About this Help System . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .311

Chapter 7 Contact Technical Support . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .313

vi NanoDrop One User Guide Thermo Scientific

1

About the NanoDrop One Spectrophotometer

The Thermo Scientific™ NanoDrop™ One is a compact,
stand-alone UV-Visible spectrophotometer developed for
micro-volume analysis of purified nucleic acids and a wide
variety of proteins. The patented sample retention system
enables the measurement of highly concentrated samples
without the need for dilutions.

The NanoDrop One system comes with preloaded software and
a touchscreen display. The instrument can be connected to an
optional USB label printer.

NOTICE Before operating a NanoDrop One instrument, please read the safety and

operating precautions and then follow their recommendations when using the instrument.

Instrument Models and Features

There are two models available for the
NanoDrop One spectrophotometer...

Optional Accessories

A number of accessories are available for the
NanoDrop One instruments...

Thermo Scientific NanoDrop One User Guide 1

Register Your Instrument

Register your instrument to receive e-mail updates on
software and...

Update Software

Quickly and easily download the latest
NanoDrop One software...

1

NanoDrop One Spectrophotometer NanoDrop OneC Spectrophotometer

Arm

Pedestal

Cuvette
holder

About the NanoDrop One Spectrophotometer

Instrument Models and Features

Instrument Models and Features

There are two models
available for the
NanoDrop One
spectrophotometer—the
NanoDrop One and the
NanoDrop One
models include the patented

micro-volume sample
retention system and general

features. The

NanoDrop One

features a cuvette holder for
analyzing dilute samples using
standard UV-visible cuvettes.

Both instruments come with a
built-in, 7-inch Android
high-resolution touchscreen
preloaded with easy-to-use
instrument control software.
The NanoDrop One software
is loaded with features to
integrate with and simplify
your daily workflows.

C

. Both

C

model also

1

Locate the instrument away from air vents and exhaust fans to minimize evaporation

2 NanoDrop One User Guide Thermo Scientific

Touchscreen

USB-A port

1

Pedestals

Touchscreen

1

About the NanoDrop One Spectrophotometer

Instrument Models and Features

Touchscreen can slide left or
right to accommodate personal
preference, and tilt forward or
back for optimal viewing

1

Two more USB-A ports are located on instrument back panel

Thermo Scientific NanoDrop One User Guide 3

1

Instrument light path

Cuvette holder

About the NanoDrop One Spectrophotometer

Instrument Models and Features

NanoDrop One Software with Acclaro Sample Intelligence Technology

The Thermo Scientific™ Acclaro™ Sample

Intelligence technology built into the

NanoDrop One instruments provides these
exclusive features to help you assess sample
integrity:

• contaminant analysis to help qualify a sample
before use in downstream applications

• on-demand technical support for
measurements that are atypical or very low
concentration

• invalid result alerts (a column sensor
monitors for the presence of bubbles or
reflective particles that can compromise
measurement results)

NanoDrop OneC Model Additional Features

C

The NanoDrop One
includes a cuvette holder for
measuring dilute samples,
colorimetric assays, cell
cultures and kinetic studies.
The cuvette system has these
additional features:

• extended lower detection

limits

• 37 °C heater option for
temperature-sensitive
samples and analyses

• micro-stirring option to
ensure sample
homogeneity and support
kinetic studies

For details, see Measure a

Sample using a Cuvette.

model

4 NanoDrop One User Guide Thermo Scientific

Optional Accessories

A number of accessories are available for the NanoDrop One instruments. To order an
accessory, contact your local distributor or visit our website.

DYMO™ LabelWriter™ 450 USB Label Printer

Prints two 5/16-in x 4-in self-adhesive labels for transferring sample data directly into
laboratory notebooks or posting on bulletin boards. The software allows printing of data from
each sample measurement or from a group of samples logged and measured together.

The printer connects to the instrument (front or back) via a USB cable (included).

PR-1 Pedestal Reconditioning Kit

1

About the NanoDrop One Spectrophotometer

Optional Accessories

Specially formulated conditioning compound that can be
applied to the pedestals to restore them to a hydrophobic state
(required to achieve adequate surface tension for accurate
sample measurements). The kit includes conditioning
compound and applicators. For more information, see

Reconditioning the Pedestals.

PV-1 Performance Verification Solution

Liquid photometric standard used to check instrument performance. For more information,
see Performance Verification.

Thermo Scientific NanoDrop One User Guide 5

1

About the NanoDrop One Spectrophotometer

Register Your Instrument

Register Your Instrument

Register your instrument to receive e-mail updates on software and accessories
for the NanoDrop One instruments. An Internet connection is required for
registration.

 To register your instrument

1. Do one of the following:

–From the NanoDrop One Viewer software running on a personal

computer (PC) that is connected to the Internet, open the Help menu
and choose NanoDrop One Website.

– From any PC that is connected to the Internet, use any web browser to

navigate to our website.

2. On the website, locate NanoDrop One Registration and follow the
instructions to register the instrument.

6 NanoDrop One User Guide Thermo Scientific

Update Software

1

About the NanoDrop One Spectrophotometer

Quickly and easily download and install the latest NanoDrop One software and
release notes from our website. Follow the steps to update or upgrade the software
on your local instrument and/or install or update the NanoDrop One Viewer
software on a personal computer (PC). An Internet connection is required to
download software.

Update Software

To install or update NanoDrop One Viewer software

1. Do one of the following:

• To install the Viewer software on a computer for the first time, open any

web browser and find the NanoDrop website.

• To update or upgrade the Viewer software, from the Viewer Home screen,

open the Help menu and choose NanoDrop One Website to open our
website.

2. On the NanoDrop website, locate the software downloads page.

3. Select to download NanoDrop One (PC) Viewer software (English version)
and follow the instructions to download and run the installer. (A computer
restart is required after the installer completes.)

4. To add a language, including software and Help systems, download and run the
language pack installer (English must be installed first). (No computer restart is
required after a language installer completes.)

Thermo Scientific NanoDrop One User Guide 7

1

About the NanoDrop One Spectrophotometer

Update Software

To update or upgrade NanoDrop One instrument software

1. Do one of the following:

2. Insert a USB device such as a memory stick into a USB port on the computer.

3. On the NanoDrop website, locate the software downloads page, select to

4. To add a language, including software and Help systems, download the

5. Insert the USB device into any USB port on the NanoDrop One instrument.

–From the NanoDrop One Viewer software, open the Help menu and

choose NanoDrop One Website to open our website.

– From any personal computer that is connected to the Internet, navigate to

the NanoDrop website.

update or upgrade NanoDrop One operating software (English version) and
follow the instructions to download the installer to the USB device.

language pack installer(s) to the USB device.

6. From the instrument Home screen, tap (Settings) > System > Update
Software.

If the USB device contains more than one version of the installer, a message is
displayed. Select the version to install (English installer must be run first) and
tap Update. (An instrument restart is required after the English installer
completes.)

When the installation is complete, a message similar to the following appears
next to the Update Software button:

Version: 1.2.0 (currently installed version of instrument operating software)
Database version: 1 (version of NanoDrop One database on this instrument)

7. To add a language, including software and Help systems, tap Update Software
again, select the language and version to install and tap Update. (No
instrument restart is required after a language installer completes.)

Note: To change the language, tap Language, select an installed language and tap
OK. (An instrument restart is required after you change the language.)

8 NanoDrop One User Guide Thermo Scientific

2

Applications

Detection Limits for All Applications

Note Detection limits provided in the tables below are approximate and apply to

micro-volume measurements only; they are based on the instrument’s photometric
absorbance range (10 mm equivalent) of 0–550 A. For measurements with 10 mm
pathlength cuvettes, the photometric absorbance range is 0–1.5 A.

Detection limits for standard applications

Sample Type Lower Detection Limit Upper Detection Limit Typical Reproducibility

dsDNA 2.0 ng/μL (pedestal)

0.20 ng/μL (cuvette)

ssDNA 1.3 ng/μL (pedestal)

0.13 ng/μL (cuvette)

RNA 1.6 ng/μL (pedestal)

0.16 ng/μL (cuvette)

Thermo Scientific NanoDrop One User Guide 9

27,500 ng/μL (pedestal)

75 ng/μL (cuvette)

18,150 ng/μL (pedestal)

49.5 ng/μL (cuvette)

22,000 ng/μL (pedestal)

60 ng/μL (cuvette)

±2.0 ng/μL for sample concentrations
between 2.0 and 100 ng/μL samples;
±2% for samples >100 ng/μL

±2.0 ng/μL for sample concentrations
between 2.0 and 100 ng/μL samples;
±2% for samples >100 ng/μL

±2.0 ng/μL for sample concentrations
between 2.0 and 100 ng/μL samples;
±2% for samples >100 ng/μL

a

2

Applications

Detection Limits for All Applications

Sample Type Lower Detection Limit Upper Detection Limit Typical Reproducibility

DNA Microarray
(ssDNA)

Purified BSA by
Protein A280

IgG by Protein
A280

Purified BSA by

1.3 ng/μL (pedestal)

0.13 ng/μL (cuvette)

0.06 mg/mL (pedestal)

0.006 mg/mL (cuvette)

0.03 mg/mL (pedestal)

0.003 mg/mL (cuvette)

0.06 mg/mL (pedestal)

495 ng/μL (pedestal)

±2.0 ng/μL for sample concentrations
between 2.0 and 100 ng/μL samples;

49.5 ng/μL (cuvette)

825 mg/mL (pedestal)

±2% for samples >100 ng/μL

±0.10 mg/mL (for 0.10–10 mg/mL samples);
±2% for samples >10 mg/mL

402 mg/mL (pedestal)

19 mg/mL (pedestal) ±0.10 mg/mL for 0.10–10 mg/mL samples

Proteins & Labels

0.006 mg/mL (cuvette)

Protein BCA 0.2 mg/mL (20:1

8.0 mg/mL (pedestal)

2% over entire range

reagent/sample volume)

0.20 mg/mL (cuvette)

0.01 mg/mL over entire range

0.01 mg/mL (1:1
reagent/sample volume)

Protein Lowry 0.2 mg/mL (pedestal) 4.0 mg/mL (pedestal) 2% over entire range

Protein Bradford 100 μg/mL (50:1

reagent/sample volume)

8000 μg/mL

±25 μg/mL for 100–500 μg/mL samples
±5% for 500–8000 μg/mL samples

a

15 μg/mL (1:1
reagent/sample volume)

Protein Pierce 660 50 μg/mL (15:1

reagent/sample volume)

25 μg/mL (7.5:1
reagent/sample volume)

a

Based on five replicates (SD=ng/μL; CV=%)

Note To minimize instrument error with highly concentrated samples, make dilutions to
ensure that measurements are made within these absorbance limits:

• For micro-volume measurements, maximum absorbance at 260 nm (for nucleic acids)
or 280 nm (for proteins) should be less than 62.5 A.

• For measurements with 10 mm pathlength cuvettes, maximum absorbance at 260 nm
(or 280 nm for proteins) should be less than 1.5 A, which is approximately 75 ng/μL
dsDNA.

100 μg/μL

2000 μg/mL

1000 μg/mL

±4 μg/mL for 15–50 μg/mL samples
±5% for 50–125 μg/mL samples

±3 μg/mL for 50–125 μg/mL samples
±2% for samples > 125 μg/mL

±3 μg/mL for 25–125 μg/mL samples
±2% for samples >125 μg/mL

10 NanoDrop One User Guide Thermo Scientific

Detection limits for pre-defined dyes

2

Applications

Detection Limits for All Applications

Sample Type Lower Detection Limit Upper Detection Limit

Cy3, Cy3.5, Alexa Fluor

0.2 pmol/μL (pedestal) 100 pmol/μL (pedestal) ±0.20 pmol/μL for sample

555, Alexa Fluor 660

a

Typical Reproducibility

concentrations between 0.20 and 4.0

b

pmol/μL;
±2% for samples >4.0 pmol/μL

Cy5, Cy5.5, Alexa Fluor
647

0.12 pmol/μL (pedestal) 60 pmol/μL (pedestal) ±0.12 pmol/μL for sample concentrations

between 0.12 and 2.4 pmol/μL;
±2% for samples >2.4 pmol/μL

Alexa Fluor 488, Alexa
Fluor 594

0.4 pmol/μL (pedestal) 215 pmol/μL (pedestal) ±0.40 pmol/μL for sample concentrations

between 0.40 and 8.0 pmol/μL;
±2% for samples >8.0 pmol/μL

Alexa Fluor 546 0.3 pmol/μL (pedestal) 145 pmol/μL (pedestal) ±0.30 pmol/μL for sample concentrations

between 0.30 and 6.0 pmol/μL;
±2% for samples >6.0 pmol/μL

a

Values are approximate

b

Based on five replicates (SD=ng/μL; CV=%)

Thermo Scientific NanoDrop One User Guide 11

Measure dsDNA, ssDNA or RNA

Measures the concentration of
purified dsDNA, ssDNA or RNA
samples that absorb at 260 nm.

Measure dsDNA, ssDNA or RNA

Reported Results

Settings

Detection Limits

Calculations

Measure dsDNA, ssDNA or RNA

Use the dsDNA, ssDNA and RNA applications to quantify purified double-stranded (ds) or
single-stranded (ss) DNA or RNA samples. These applications report nucleic acid
concentration and two absorbance ratios (A260/A280 and A260/A230). A single-point
baseline correction can also be used.

To measure dsDNA, ssDNA or RNA samples

NOTICE

• Do not use a squirt or spray bottle on or near the instrument as liquids will flow into
the instrument and may cause permanent damage.

• Do not use hydrofluoric acid (HF) on the pedestals. Fluoride ions will permanently
damage the quartz fiber optic cables.

Thermo Scientific NanoDrop One User Guide 13

Measure dsDNA, ssDNA or RNA

Typical nucleic acid spectrum

Comparison of nucleic acid spectra with
and without two common contaminants

Before you begin...

Before taking pedestal measurements with the NanoDrop One instrument, lift the instrument
arm and clean the upper and lower pedestals. At a minimum, wipe the pedestals with a new
laboratory wipe. For more information, see Cleaning the Pedestals.

 To measure nucleic acid

1. From the Home screen, select the Nucleic Acids tab
and tap dsDNA, ssDNA or RNA, depending on the
samples to be measured.

2. Specify a baseline correction if desired.

3. Pipette 1–2 μL blanking solution onto the lower
pedestal and lower the arm, or insert the blanking
cuvette into the cuvette holder.

Tip: If using a cuvette, make sure to align the cuvette

light path with the instrument light path.

4. Tap Blank and wait for the measurement to
complete.

Tip: If Auto-Blank is On, the blank measurement
starts automatically after you lower the arm. (This
option is not available for cuvette measurements.)

5. Lift the arm and clean both pedestals with a new
laboratory wipe, or remove the blanking cuvette.

6. Pipette 1-2 μL sample solution onto the pedestal and
lower the arm, or insert the sample cuvette into the
cuvette holder.

7. Start the sample measurement:

–Pedestal: If Auto-Measure is On, lower arm; if

Auto-Measure is off, lower arm and tap Measure.

– Cuvette: Tap Measure.

When the sample measurement is completed, the
spectrum and reported values are displayed (see the
next section).

8. When you are finished measuring samples, tap
End Experiment.

9. Lift the arm and clean both pedestals with a new
wipe, or remove the sample cuvette.

14 NanoDrop One User Guide Thermo Scientific

Measure dsDNA, ssDNA or RNA

Best practices for nucleic acid measurements

• Isolate and purify nucleic acid samples before measurement to remove impurities.
Depending on the sample, impurities could include DNA, RNA, free nucleotides,
proteins, some buffer components and dyes. See Preparing Samples for more information.

Note Extraction reagents such as guanidine, phenol, and EDTA contribute
absorbance between 230 nm and 280 nm and will affect measurement results if
present in samples (even residual amounts).

• Ensure the sample absorbance is within the instrument’s absorbance detection limits.

• Blank with the same buffer solution used to resuspend the analyte of interest. The
blanking solution should be a similar pH and ionic strength as the analyte solution.

•Run a blanking cycle to assess the absorbance contribution of your buffer solution. If the
buffer exhibits strong absorbance at or near the analysis wavelength (typically 260 nm),
you may need to choose a different buffer or application. See Choosing and Measuring a

Blank for more information.

• For micro-volume measurements:

– Ensure pedestal surfaces are properly cleaned and conditioned.

– If possible, heat highly concentrated or large molecule samples, such as genomic or

lambda DNA, to 63 °C (145 °F) and gently (but thoroughly) vortex before taking a
measurement. Avoid introducing bubbles when mixing and pipetting.

– Follow best practices for micro-volume measurements.

– Use a 1-2 μL sample volume. See Recommended Sample Volumes for more

information.

C

• For cuvette measurements (NanoDrop One

instruments only), use compatible cuvettes

and follow best practices for cuvette measurements.

Related Topics

• Measure a Micro-Volume Sample

• Measure a Sample Using a Cuvette

• Best Practices for Micro-Volume Measurements

• Best Practices for Cuvette Measurements

• Prepare Samples and Blanks

• Basic Instrument Operations

Thermo Scientific NanoDrop One User Guide 15

Measure dsDNA, ssDNA or RNA

Tap row to
select sample
and update
spectrum; tap
more rows to
overlay up to five
spectra. Press
and hold
sample row to
view
measurement
details.

Drag tab
down/up to see

more/less
sample data

Nucleic acid
concentration

UV spectrum

Tap to select unit

Purity ratios

Menu of options;
tap to open

Sample name;

tap to edit

Swipe screen left to

view table with more
measurement results

Pinch and zoom to
adjust axes;
double-tap to reset

Tap to end
experiment and
export data

Menu of options;

tap to open

Nucleic Acid Reported Results

dsDNA measurement screen

For each measured sample, the dsDNA, ssDNA and RNA applications show the UV
absorbance spectrum and a summary of the results. Here is an example:

Note Micro-volume absorbance measurements and measurements taken with
nonstandard cuvettes are normalized to a 10.0 mm pathlength equivalent.

16 NanoDrop One User Guide Thermo Scientific

Measure dsDNA, ssDNA or RNA

dsDNA, ssDNA and RNA reported values

The initial screen that appears after each measurement (see previous image) shows a summary
of the reported values. To view all reported values, press and hold the sample row. Here is an
example:

• sample details (application and sampling method used, i.e., pedestal or cuvette)

•sample name

• created on (date sample measurement was taken)

• nucleic acid concentration

• A260/A280

• A260/A230

•A260

•A280

•factor

• baseline correction

Related Topics

• Basic Instrument Operations

• Nucleic Acid Calculations

Setting for Nucleic Acid Measurements

To show the dsDNA, ssDNA or RNA settings, from the dsDNA, ssDNA or RNA
measurement screen, tap > Nucleic Acid Setup.

Thermo Scientific NanoDrop One User Guide 17

Measure dsDNA, ssDNA or RNA

Setting Available Options Description

Baseline Correction On or off

Enter baseline correction
wavelength in nm or use
default value (340 nm)

Optional user-defined baseline correction. Can be used to
correct for any offset caused by light scattering particulates by
subtracting measured absorbance at specified baseline correction
wavelength from absorbance values at all wavelengths in sample
spectrum. As a result, absorbance of sample spectrum is zero at
specified baseline correction wavelength.

Related Topics

• Instrument Settings

Calculations for Nucleic Acid Measurements

The nucleic acid applications use the Beer-Lambert

equation to correlate absorbance with concentration.

Solving Beer’s law for concentration yields the equation
at the right.

Beer-Lambert Equation (solved for concentration)

c = A / ( * b)

where:

A = UV absorbance in absorbance units (AU)

 = wavelength-dependent molar absorptivity coefficient (or extinction

coefficient) in liter/mol-cm

b = pathlength in cm

c = analyte concentration in moles/liter or molarity (M)

Note: Dividing the measured absorbance of a sample solution by its molar
extinction coefficient yields the molar concentration of the sample. See

Published Extinction Coefficients for more information regarding molar

vs. mass concentration values.

18 NanoDrop One User Guide Thermo Scientific

Measure dsDNA, ssDNA or RNA

The Nucleic Acid applications use a modification of the
Beer-Lambert equation (shown at right) to calculate
sample concentration where the extinction coefficient
and pathlength are combined and referred to as a
“factor.”

For the dsDNA, ssDNA and RNA applications, the
generally accepted factors for nucleic acids are used in
conjunction with Beer’s Law to calculate sample
concentration. For the Custom Factor application, the
user-specified factor is used.

Extinction Coefficients vs Factors

Using the terms in the Beer-Lambert equation, factor (f) is defined as:

factor (f) = 1/( * b)

where:

 = wavelength-dependent molar extinction coefficient in ng-cm/μL

b = sample pathlength in cm

As a result, analyte concentration (c) is calculated as:

c = A * [1/( * b)]

or

c = A * f

where:

c = analyte concentration in ng/μL
A = absorbance in absorbance units (A)

f = factor in ng-cm/μL (see below)

Factors Used

• dsDNA (factor = 50 ng-cm/μL)

• ssDNA (factor = 33 ng-cm/μL)

• RNA (factor = 40 ng-cm/μL)

• Custom Factor (user entered factor between 15 ng-cm/μL and
150 ng-cm/μL

Thermo Scientific NanoDrop One User Guide 19

Measure dsDNA, ssDNA or RNA

Calculated nucleic acid concentrations are based on the
absorbance value at 260 nm, the factor used and the
sample pathlength. A single-point baseline correction (or
analysis correction) may also be applied.

Concentration is reported in mass units. Calculators are
available on the Internet to convert concentration from
mass to molar units based on sample sequence.

Absorbance values at 260 nm, 280 nm and sometimes
230 nm are used to calculate purity ratios for the
measured nucleic acid samples. Purity ratios are
sensitive to the presence of contaminants in the sample,
such as residual solvents and reagents typically used
during sample purification.

Measured Values

Note: For micro-volume absorbance measurements and measurements

taken with nonstandard (other than 10 mm) cuvettes, the spectra are
normalized to a 10 mm pathlength equivalent.

A260 absorbance

• Nucleic acid absorbance values are measured at 260 nm using the
normalized spectrum. This is the reported A260 value if Baseline
Correction is not selected.

•If Baseline Correction is selected, the absorbance value at the
correction wavelength is subtracted from the absorbance at 260 nm.
The corrected absorbance at 260 nm is reported and used to calculate
nucleic acid concentration.

A230 and A280 absorbance

• Normalized and baseline-corrected (if selected) absorbance values at
230 nm and 280 nm are used to calculate A260/A230 and A260/A280
ratios.

Sample Pathlength

• For micro-volume measurements, the software selects the optimal
pathlength (between 1.0 mm and 0.03 mm) based on sample
absorbance at the analysis wavelength.

• For cuvette measurements, pathlength is determined by the cuvette
Pathlength setting in the software (see General Settings).

• Displayed spectra and absorbance values are normalized to a 10 mm
pathlength equivalent.

20 NanoDrop One User Guide Thermo Scientific

Measure dsDNA, ssDNA or RNA

Reported Values

• Nucleic acid concentration. Reported in selected unit (i.e., ng/μL,
μg/uL or μg/mL). Calculations are based on modified Beer’s Law
equation using corrected nucleic acid absorbance value.

• A260/A280 purity ratio. Ratio of corrected absorbance at 260 nm to
corrected absorbance at 280 nm. An A260/A280 purity ratio of ~1.8 is
generally accepted as “pure” for DNA (~2.0 for RNA). Acidic solutions
may under represent the reported value by 0.2-0.3; the opposite is true
for basic solutions.

• A260/A230 purity ratio. Ratio of corrected absorbance at 260 nm to
corrected absorbance at 230 nm. An A260/A230 purity ratio between

1.8 and 2.2 is generally accepted as “pure” for DNA and RNA.

Note: Although purity ratios are important indicators of sample quality,
the best quality indicator quality is functionality in the downstream
application of interest (e.g., real-time PCR).

Thermo Scientific NanoDrop One User Guide 21

Measure Microarray

Measures the concentration of
purified nucleic acids that have been
labeled with up to two fluorescent
dyes for use in downstream
microarray applications.

Measure Microarray Samples

Reported Results

Settings

Detection Limits

Calculations

Measure Microarray Samples

Use the Microarray application to quantify nucleic acids that have been labeled with up to
two fluorescent dyes. The application reports nucleic acid concentration, an A260/A280 ratio
and the concentrations and measured absorbance values of the dye(s), allowing detection of
dye concentrations as low as 0.2 picomole per microliter.

To measure microarray samples

NOTICE

• Do not use a squirt or spray bottle on or near the instrument as liquids will flow into
the instrument and may cause permanent damage.

• Do not use hydrofluoric acid (HF) on the pedestals. Fluoride ions will permanently
damage the quartz fiber optic cables.

Thermo Scientific NanoDrop One User Guide 23

Measure Microarray

Before you begin...

Before taking pedestal measurements with the NanoDrop One instrument, lift the instrument
arm and clean the upper and lower pedestals. At a minimum, wipe the pedestals with a new
laboratory wipe. For more information, see Cleaning the Pedestals.

24 NanoDrop One User Guide Thermo Scientific