3 Kit SMART Digest 15 digestion - Thermo Fisher Scientific 516.70 516.75 516.80 516.85 516.90 Mass...
1
Peptide Mapping Sample to knowledge easier and faster than before Peptide maps require 100% sequence coverage to prove the molecular structure of the biotherapeutic Digestion Separation Rituximab heavy chain Automation For more information: thermofisher.com/SMARTdigest For more information on peptide mapping: thermofisher.com/peptidemaps For technical information: thermofisher.com/chromexpert 5 10 15 20 25 30 0 Minutes mAU 200 RSD (n=3, 15 peaks) Retention Time: 0.024% Peak Area: 4.466% n=3 RT RSD (%) Less than retain 100% sequence coverage Thermo Scientific ™ BioPharma Finder ™ software allows easy interrogation of peptide maps enabling full characterization and confidence in results ©2017 Thermo Fisher Scientific. All rights reserved. XX21695-EN 0517S Overlay of 3 separate digestions of a mAb using the SMART Digest kit (15mins), by 3 individual users Rituximab light chain Easy SMART Digest Kit 3 Digestion with the Steps SMART Digestion Add sample Digestion Add buffer 1400 RPM 70 oC In-solution digestion 15 Steps Faster Reproducibility Typical Digestion Times Protein Digest Time (min) Insulin 4 BSA < 5 Carbonic anhydrase < 5 Lysozyme < 5 Apo-B 30 IgG 45 IgG in 50 µL plasma* 75 Ribonuclease A 150 Thyroglobulin 240 C-reactive protein 240 200 μL protein solution (100 μg/mL) at 70 °C *IgG in plasma (17.5 mg/mL total protein) at 70 °C Using the Thermo Scientific ™ SMART Digest ™ kit thyroglobulin is digested in 3.5 hours versus 20 hours with in-solution digestion Faster 6x 1 2 3 The SMART Digest kit is highly reproducible user-to-user. Peak area RSD of... < 5% Automate 96 Digestion of samples can be easily automated with the Thermo Scientific ™ KingFisher ™ purification systems, allowing for fast, high throughput sample processing. KingFisher systems help you to reduce hands on time and bring reproducibility to your workflow. Samples Easy mAU 2.5 4 31.3 -45.2 Min 0.0 201.0 25.0 50.0 75.0 100.0 125.0 150.0 175.0 -25.0 6 8 10 12 14 16 18 20 22 24 26 28 30 Mass Detection Highly reproducible Faster separation High mass accuracy in Peptide Confidence Mapping HRAM MS +/- 3 ppm Intensity (%) Wrong answer for both peptides Right answer for both peptides 0 20 40 60 80 100 516.65 516.70 516.75 516.80 516.85 516.90 Mass (amu) 0 20 40 60 80 100 516.65 516.70 516.75 516.80 516.85 516.90 Mass (amu) 516.78490 (correct) 516.76671 (correct) 516.77581 (observed) Intensity (%) 516.78490 516.76671 R = 15,000 R = 56,700 Confidence in results Data Interpretation Peptide mixture: [Val5]-Angiotensin II [Lys-des-Arg 9 ]-Bradykinin Sequence: DRVYVHPF KRPPGFSPF Formula: C 49 H 69 N 13 O 12 C 50 H 73 N 13 O 11 Exact mass: [M+2H] 2+ = 516.76671 [M+2H] 2+ = 516.78490 Dm (mmu): 18.2 mmu Joshua J. Coon, et al. ASMS 2012 oral, MOB pm Easy data interpretation + Overlay of 13 consecutive chromatograms of a mAb digested using the SMART Digest kit Achieve Retention time repeatability with the Thermo Scientific ™ Vanquish ™ Horizon UHPLC system Faster run times
Transcript of 3 Kit SMART Digest 15 digestion - Thermo Fisher Scientific 516.70 516.75 516.80 516.85 516.90 Mass...
Peptide Mapping Sample to knowledge easier and faster than beforePeptide maps require 100% sequence coverage to prove the molecular structure of the biotherapeutic
Digestion
Separation
Rituximab heavy chain
Automation
For more information: thermofisher.com/SMARTdigest For more information on peptide mapping: thermofisher.com/peptidemapsFor technical information: thermofisher.com/chromexpert
5 10 15 20 25 30
0
Minutes
mAU
200
RSD (n=3, 15 peaks)Retention Time: 0.024%Peak Area: 4.466%
n=3
RT RSD (%) Less than
retain 100% sequence coverage
Thermo Scientific™ BioPharma Finder™ software allows easy interrogation of peptide maps enabling full characterization and confidence in results
©2017 Thermo Fisher Scientific. All rights reserved. XX21695-EN 0517S
Overlay of 3 separate digestions of a mAb using the SMART Digest kit (15mins), by 3 individual users
Rituximab light chain
Easy
SMART Digest Kit3
Digestion with the
Steps
SMART Digestion
Add sample DigestionAdd buffer
1400 RPM 70 oC
In-solution digestion
15Steps
Faster
Reproducibility
Typical Digestion Times
Protein Digest Time (min)
Insulin 4
BSA < 5
Carbonic anhydrase < 5
Lysozyme < 5
Apo-B 30
IgG 45
IgG in 50 µL plasma* 75
Ribonuclease A 150
Thyroglobulin 240
C-reactive protein 240
200 μL protein solution (100 μg/mL) at 70 °C*IgG in plasma (17.5 mg/mL total protein) at 70 °C
Using the Thermo Scientific™ SMART Digest™ kit thyroglobulin is digested in 3.5 hours versus 20 hours with in-solution digestion
Faster6x
1 2 3
The SMART Digest kit is highly reproducible user-to-user.Peak area RSD of...
< 5%
Automate
96Digestion of samples can be easily automated with the Thermo Scientific™ KingFisher™ purification systems, allowing for fast, high throughput sample processing. KingFisher systems help you to reduce hands on time and bring reproducibility to your workflow.
Samples
Easy
mAU
2.5 4 31.3-45.2
Min
0.0
201.0
25.0
50.0
75.0
100.0
125.0
150.0
175.0
-25.0
6 8 10 12 14 16 18 20 22 24 26 28 30
Mass Detection
Highly reproducible
Faster separation
High mass accuracy
in Peptide Confidence
MappingHRAM MS+/- 3 ppm
Inten
sity (
%)
Wrong answer for both peptides Right answer for both peptides
0
20
40
60
80
100
516.65 516.70 516.75 516.80 516.85 516.90
Mass (amu)
0
20
40
60
80
100
516.65 516.70 516.75 516.80 516.85 516.90
Mass (amu)
516.78490(correct)
516.76671(correct)
516.77581 (observed)
Inten
sity (
%)
516.78490516.76671
R = 15,000 R = 56,700
Confidence in results
Data Interpretation
Peptide mixture: [Val5]-Angiotensin II [Lys-des-Arg9]-BradykininSequence: DRVYVHPF KRPPGFSPF Formula: C
49H
69N
13O
12 C
50H
73N
13O
11
Exact mass: [M+2H]2+ = 516.76671 [M+2H]2+ = 516.78490
Dm (mmu): 18.2 mmuJoshua J. Coon, et al. ASMS 2012 oral, MOB pm
Easy data interpretation
+
Overlay of 13 consecutive chromatograms of a mAb digested using the SMART Digest kit
Achieve
Retention time repeatability with the Thermo Scientific™ Vanquish™ Horizon UHPLC system
Fasterrun times
