（M） Flow rate Detector Column temp. : Room temp.342112 () ili Aqueous SECGFCCoumns: 13452312132544123531231205056872015 005553213505 505lSca-based(A) LW-403 4D（D）（M）(B) LW-803Column Eluent : (A) Shodex PROTEIN LW-403 4D (B) Shodex PROTEIN LW-803: 50mM Sodium phosphate buffer (pH7.0) + 0.3M NaCl: (A) 0.35 mL/min (B) 1.0 mL/min: (A) UV (280 nm) (small cell volume) (B) UV (280 nm) (conventional type)Column Eluent Flow rate Detector Column temp. : 30 ˚C: Shodex KW402.5-4F: 50 mM Sodium phosphate buffer (pH7.0) + 0.3 M NaCl: 0.33 mL/min: UV (220 nm) (small cell volume)PROTEIN LW-803 is suitable for analyzing a few-hundred-thousand molecular weight size proteins. When comparing LW-803 to our conventional columns and other manufacturer's columns, LW-803 provides a better separation around 160,000 molecular weight range that is about the size of Globulin. This improved separation ef■ciency is advantageous for the separation of monomer and dimer of IgG which is a mainstream of antibody drug.Sample : 5 µL 1. Thyroglobulin 2. γ-Globulin 3. Ovalbumin 4. Ribonuclease A (MW : 13,700) 7 mg/mL5. Uridine LW-803KW-803Company A: Shodex PROTEIN LW-803, Shodex PROTEIN KW-803, Silica-based SEC column from other manufacturerColumn : 50 mM Sodium phosphate buffer (pH7.0) + 0.3 M NaClEluent : 1.0 mL/minFlow rate Detector : UV (280 nm)Column temp. : Room temp.Papain digestion of humanized monoclonal IgG was monitored using PROTEIN LW-803, an aqueous SEC (GFC) column. During the papain digestion of IgG, Fc and Fab fragments from the IgG and their decomposition intermediates are expected to be observed. LW-803 separates IgG and decomposed fragments and intermediates well from each other, thus it is suitable for the monitoring of papain digestion of IgG.Papain digestion(24 hr)Papain digestion(12 hr)Papain digestion(4 hr)Papain digestion (90 min)Papain digestion (30 min)Papain digestion (0 min)Without PapainKW400 series is a high performance type semi-micro columns. It offers approximately 1.5 times larger theoretical plate number and 3 to 4 times higher detection sensitivity (peak height) than KW-800 series columns do.Sample : 10 µL1. Blue dextran 2000 0.2 mg/mL0.8 mg/mL2. γ-Globulin 0.8 mg/mL3. Ovalbumin 4. Myoglobin 0.56 mg/mL0.04 mg/mL5. Uridine （A）KW402.5-4F（B）KW-802.5Column Eluent Flow rate Detector Column temp. : 25 ˚C: (A) Shodex KW402.5-4F (B) Shodex PROTEIN KW-802.5: 50 mM Sodium phosphate buffer (pH7.0) + 0.3 M NaCl : (A) 0.33 mL/min, (B) 1.0 mL/min : UV (280 nm) (small cell volume)(MW : 670,000) 7 mg/mL(MW : 160,000) 6 mg/mL(MW : 44,300) 4.8 mg/mL(MW : 244) 0.1 mg/mL(conventional type)345TP : 17,0004TP : 8,2004(Procedures for digestion monitoring)(1) Dissolve 3 mg of humanized monoclonal IgG in 500 µL of the eluent. (6 mg/mL conc.)(2) Dissolve 1 mg of papain in 500 µL of the eluent. (1 mg/mL conc.)(3) Filer (1) and (2) using 0.2-µm membrane ■lters (4) Mix two solutions in 1:1 ratio. (5) Keep the mixture at 25 ºC.(6) Take samples at set timings and analyze them by HPLC.Column Eluent Flow rate Detector Column temp. : 25 ˚CColumn Eluent Flow rate Detector Column temp. : 30 ˚C10Sample : 10 µLHumanized monoclonal IgG1. Aggregates of IgG2. Dimer of IgG3. Monomer of IgG4 - 6. Fragments of IgG from papain digestion7. Citric acid8. Papain1015 minSample : Whey, 5 µL201015minSample : 5 µLIgG from human serum 10 mg/mL1. Aggregates2. Dimer3. Monomer(conventional type)15 min304050 minSample : IgG from human serum 10 mg/mL(LW-403 4D) 0.5 µL(LW-803) 5 µLAnalytical time1/2Sample : 5 µL0.6 mg/mL1. Lectin from soybean 2. Lectin from arachis hypogaea 1.1 mg/mL3. Lectin from canavalia ensiformis (Con A) 0.9 mg/mL0.7 mg/mL4. Lectin from lens culinaris (LCA) 5. Lectin from triticum vulgaris (WGA) 0.8 mg/mL1015 min(D) Dimer(M) Monomer1015min10 minLW-803KW-8032.21.6Monitoring papain digestion of humanized monoclonal IgGComparison of KW402.5-4F and KW-802.5Comparison of LW-803, conventional column, and other manufacturer's columnWhey in yogurtLW-803KW-803Company A: Shodex PROTEIN LW-803: 0.1 M Sodium phosphate buffer (pH7.0) + 0.3 M NaCl: 1.0 mL/min: UV (280 nm): Shodex KW403-4F + KW402.5-4F: 50 mM Sodium phosphate buffer (pH7.0) + 0.3 M NaCl: 0.20 mL/min: UV (280 nm) (small cell volume)Efficiencies of LW-403 4D over LW-803 for IgG separationLectinsResolution(Monomer/Dimer)1.9Company A（D）39
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