Peptide and Protein Bioanalysis: Infographic
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Peptides and proteins are not small molecules. Why treat them the same? Analyzing large molecules may be one of the greatest challenges that the bioanalyst faces. In our infographic we answer: Why is it harder to quantify large molecules? Why are biotherapeutics more important than ever? How can Waters help?
![[ PEPTIDE AND PROTEIN BIOANALYSIS ]
Aspirin 180 Da Bevacizumab
149 kDa
Insulin Glargine
6,063 Da
PEPTIDES AND PROTEINSSMALL MOLECULES
Multiple precursor
charge states dilute signal
Increased fragmentation
further reduces signal
2+
3+ 4+
5+
1+
In 2013, 7 of the top 10 best-selling prescription drugs were large molecules
2
Humira®
148 kDa
Enbrel®
150 kDa
Remicade®
149 kDa
Lantus®
6064 Da
Rituxan®
145 kDa
Avastin®
149 kDa
Herceptin®
148 kDa
Seretide®
500 Da
Crestor®
1001 Da
Abilify®
448 Da
#1 #2 #3 #4 #5 #6 #7 #8 #9 #10
Increased sensitivity and specificity challenges:
Fewer molecules in
equal volume of analyte
WHYis it harder to
quantify large molecules?
WHYare biotherapeutics
more important than ever?
HOW can Waters help?
Optimize specificity of sample prep
Oasis®
MAX and WCX SPE µElution Plates help achieve high recovery and low limits
of detection for a wide range of endogenous, therapeutic and surrogate peptides.
Wide mass range, reliable sensitivity
The Xevo®
family of tandem quadrupole mass spectrometers delivers
high sensitivity at high and low mass, detecting all fragments without
compromise, while sensitivity at high m/z enhances specificity.
Improved microfluidics for
better sensitivity and reduced sample load
The ionKey/MS™ System integrates the UPLC analytical separation directly
into the source of the mass spectrometer, delivering significant increases in
sensitivity, solvent savings, and reductions in sample volume.
Increasing sensitivity through LC optimization
$$
10x
UP TO
IMPROVEMENT IN
SENSITIVITY
More LC and MS parameters to
consider reduces efficiency
Complex sample prep means
possible loss of analyte
STEP 1 STEP 2 STEP 3 STEP 4
Year 201420102005200019951990
204
9940
1.2.801.201.00 1.00
385
568
2.402.001.201.00
MRM of 1 Channel ES+
1.32e4
Area
365
240
1.251.00 1.25
60 °C
40 °C
400 µL/min
200 µL/min
ACN + 0.1%
formic acid
ACN + 0.1%
formic acid + 5% TFE
14237
Temperature Flow rate Mobile phase B composition
CAPILLARY VOLTAGE
MOBILE PHASE COMPOSITION
CHROMATOGRAPHIC PORE SIZE
COLUMN TEMP
FLOW RATE
Analyzing large molecules may be one
of the greatest challenges that the
bioanalyst faces at the beginning
of the 21st century.
PEPTIDES AND PROTEINS
ARE NOT SMALL MOLECULES.
WHY TREAT THEM
THE SAME?
Biotherapeutics
publications1
Use a generic set of chromatographic conditions and evaluate with the ACQUITY UPLC®
BEH C18
300 Å,
ACQUITY UPLC CSH, and CORTECS®
UPLC C18
+ Columns to identify the best peak and separation and then
optimize key parameters, such as column temperature, flow rate, and mobile phase composition.
Endogenous peptides and
proteins yield high background
SENSITIVITY
SPECIFICITY
©2015 Waters Corporation 720005295EN February 2015 AW-KW
Learn more with the
Peptide Bioanalysis Solution Guide
Search literature code 720004563EN at waters.com
1
Electronic search performed in PubMed.org on
January 13, 2015 with keyword “biotherapeutics.”
2
Kollewe, J. (2014, March 27). The world's 10 best-selling
prescription drugs. The Guardian. Retrieved Jan. 13, 2015.](https://image.slidesharecdn.com/peptideandproteinbioanalysis720005295en-150211130651-conversion-gate02/85/Peptide-and-Protein-Bioanalysis-Infographic-1-320.jpg)
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Peptide and Protein Bioanalysis: Infographic
- 1. [ PEPTIDE AND PROTEIN BIOANALYSIS ] Aspirin 180 Da Bevacizumab 149 kDa Insulin Glargine 6,063 Da PEPTIDES AND PROTEINSSMALL MOLECULES Multiple precursor charge states dilute signal Increased fragmentation further reduces signal 2+ 3+ 4+ 5+ 1+ In 2013, 7 of the top 10 best-selling prescription drugs were large molecules 2 Humira® 148 kDa Enbrel® 150 kDa Remicade® 149 kDa Lantus® 6064 Da Rituxan® 145 kDa Avastin® 149 kDa Herceptin® 148 kDa Seretide® 500 Da Crestor® 1001 Da Abilify® 448 Da #1 #2 #3 #4 #5 #6 #7 #8 #9 #10 Increased sensitivity and specificity challenges: Fewer molecules in equal volume of analyte WHYis it harder to quantify large molecules? WHYare biotherapeutics more important than ever? HOW can Waters help? Optimize specificity of sample prep Oasis® MAX and WCX SPE µElution Plates help achieve high recovery and low limits of detection for a wide range of endogenous, therapeutic and surrogate peptides. Wide mass range, reliable sensitivity The Xevo® family of tandem quadrupole mass spectrometers delivers high sensitivity at high and low mass, detecting all fragments without compromise, while sensitivity at high m/z enhances specificity. Improved microfluidics for better sensitivity and reduced sample load The ionKey/MS™ System integrates the UPLC analytical separation directly into the source of the mass spectrometer, delivering significant increases in sensitivity, solvent savings, and reductions in sample volume. Increasing sensitivity through LC optimization $$ 10x UP TO IMPROVEMENT IN SENSITIVITY More LC and MS parameters to consider reduces efficiency Complex sample prep means possible loss of analyte STEP 1 STEP 2 STEP 3 STEP 4 Year 201420102005200019951990 204 9940 1.2.801.201.00 1.00 385 568 2.402.001.201.00 MRM of 1 Channel ES+ 1.32e4 Area 365 240 1.251.00 1.25 60 °C 40 °C 400 µL/min 200 µL/min ACN + 0.1% formic acid ACN + 0.1% formic acid + 5% TFE 14237 Temperature Flow rate Mobile phase B composition CAPILLARY VOLTAGE MOBILE PHASE COMPOSITION CHROMATOGRAPHIC PORE SIZE COLUMN TEMP FLOW RATE Analyzing large molecules may be one of the greatest challenges that the bioanalyst faces at the beginning of the 21st century. PEPTIDES AND PROTEINS ARE NOT SMALL MOLECULES. WHY TREAT THEM THE SAME? Biotherapeutics publications1 Use a generic set of chromatographic conditions and evaluate with the ACQUITY UPLC® BEH C18 300 Å, ACQUITY UPLC CSH, and CORTECS® UPLC C18 + Columns to identify the best peak and separation and then optimize key parameters, such as column temperature, flow rate, and mobile phase composition. Endogenous peptides and proteins yield high background SENSITIVITY SPECIFICITY ©2015 Waters Corporation 720005295EN February 2015 AW-KW Learn more with the Peptide Bioanalysis Solution Guide Search literature code 720004563EN at waters.com 1 Electronic search performed in PubMed.org on January 13, 2015 with keyword “biotherapeutics.” 2 Kollewe, J. (2014, March 27). The world's 10 best-selling prescription drugs. The Guardian. Retrieved Jan. 13, 2015.