Waters Intact mAb Mass Check Standard - Manual (Page 2)

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[ CARE AND USE MANUAL ]
Intact mAb Mass Check Standard
1
2
3
4
5
ESI-TOF
MS spectrum
Deconvoluted
TOF spectrum
+50
+47
+53
Mass (Da)
2200
147600 148000 148400 148800 149200 149600
2600 3000 3400 3800
m/z
mass
II. STORAGE AND STABILITY
The lyophilized powder is recommended to be stored long term
at -20 °C until the expiration date printed on the label. Once
reconstituted, the solution is stable for one week when stored at 8 °C.
III. RECONSTITUTION OF THE INTACT mAB MASS
CHECK STANDARD FOR MS OPTIMIZATION
The Intact mAb Mass Check Standard is 1 mg per vial of a
lyophilized solid. Depending on system requirements please
follow the below solubilization instructions:
SYNAPT G1/G2 and Xevo G1/G2
Add 100 µL of DI water to the Max Recovery Vial. Sonicate for 5
minutes. The mAb concentration of this solution will be 10 mg/
mL or 67 µM. Dilute this solution 20 fold with Eluent A of the LC
mobile phase to prepare a solution containing 3.3 µM mAb and
inject 5-20 µL for testing the LC/MS system.
SYNAPT/Xevo G2-S
Add 100 µL of DI water to the Max Recovery Vial. Sonicate for
5 minutes. The mAb concentration of this solution will be 10 mg/mL
or 67 µM. Dilute this solution 200 fold with Eluent A of the LC
mobile phase to prepare a solution containing 0.33 µM mAb
and inject 5-20 µL for testing the LC/MS system.
SQD
Add 200 µL of DI water to the Max Recovery Vial. Sonicate for
5 minutes. The mAb concentration of this solution will be 5 mg/
mL or 33 µM. Inject 5-10 µL for testing the LC/MS system.
IV. MOLECULAR WEIGHT INFORMATION
Light Chain Formula: Avg. MW =
C
1066
H
1649
N
287
O
343
S
7
24198 Da
Heavy Chain Formula: Avg. MW =
C
2170
H
3338
N
562
O
661
S
19
484848 Da
Intact Protein Formula: Average MW =
C
6472
H
9940
N
1698
O
2008
S
52
145,330 Da
Total number of Disulfide Bond = 17
Fixed Modification on 2 x Pyroglutamic Acid Q
For sequence information to cut/paste please go to:
http://www.waters.com/waters/nav.htm?cid=134634380 and
Click on Intact mAb Mass Check Standard.
V. EXAMPLES OF USING THE INTACT mAB
MASS CHECK STANDARD FOR MS
OPTIMIZATION WITH Q-Tof, Tof AND SQD
Table 1: Deconvoluted MW of the Major Glycoform of the mAb
Peak Number mAb Glycoform Expected MW
1 M + G0F + G0F 148,220
2 M + G0F + G1F 148,383
3 M + G1F + G1F 148,545
4 M + G1F + G2F 148,707
5 M + G2F + G2F 148,868
*aglycosylated mAb
Figure 1: Example Data obtained with ESI-TOF Instrumentation.