UHPLC Analysis of Orthoclone OKT3 IgG2 Monoclonal Antibody on BIOshell A400 Protein C4
CONDITIONS
column
BIOshell A400 Protein C4; 10 cm x 2.1 mm I.D., 3.4 μm particles (66825-U)
mobile phase
[A] 90:10 (0.1% TFA in water): (0.1% TFA in acetonitrile); [B] 10:90 (0.1% TFA in water): (0.1% TFA in acetonitrile)
gradient
0% B to 50%B in 50 min; 50%B to 100% B in 10 min
flow rate
0.2 mL/min
pressure
1650 psi (114 bar)
column temp.
75 °C
detector
UV, 215 nm
injection
1.0 μL
sample
Orthoclone OKT3, 1 mg/mL, 0.05% TFA in water
Description
Analysis Note
Orthoclone OKT3 (common name: muromonab-CD3) is a recombinant monoclonal antibody (mAb) to human CD3. This mAb is used as an immunosuppressant drug given to patients to reduce acute rejection of foreign organs during an organ transplant. This mAb also has the distinction of being the first mAb to be approved for clinical use in humans by the FDA. Due to its use as a therapeutic, the finished drug product needs to be carefully characterized in order to make sure that it is impurity and/or degradant free. One way to characterize these therapeutic mAbs is by reversed-phase chromatography (RPC). A RPC method was developed that allows for resolution of the main mAb peak from impurities present in the sample. The inset shows the region around the main analyte, and gives an idea on how complex therapeutic mAb samples can be.
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