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1-1 UHPLC-SEC 法对治疗性抗体的分析 TSKgel UP-SW3 UHPLC HPLC A TSKgel UP-SW3 2 μm (4.6 mm ID. 15 cm B UHPLC-SEC 1.7 μm (4.6 mm ID. 15 cm mmol/l (ph 6.7 + 2SO4 mmol/l Na +.5 % NaN3.35 ml/ 25 UV 28 nm 5 μl - IgG 1. 2. 3. 4. 1-2 UHPLC-SEC 法对治疗性抗体的高分辨率 快速分析 UP-SW3 2 G3SWXL UP-SW3 G3SWXL A TSKgel G3SW XL 5 μm (7.8 mm ID. 3 cm 2 B TSKgel UP-SW3 2 μm (4.6 mm ID. 3 cm mmol/l (ph 6.7 + mmol/l Na2SO4 +.5 % NaN3 A 1. ml/ B.35 ml/ 25 UV 28 nm μl - IgG 1. 2. 3. 4. ~ 3 ~
1-3 SEC 法对抗体药物的超快速分析 TSKgel SuperSW mab HTP HPLC :.5mL/ : 5.MPa Rs / 1.91 :.35mL/ : 3.6MPa R s / 2.13 TSKgel SuperSW mab HTP (4.6 mm ID 15 cm.2 mol/l (ph 6.7 +.5% NaN3.5 ml/(.35 ml/( UV 28 nm 25 μl - IgG 1 2 3 4 5 6 7 1-4 SEC 法对抗体多聚体 ( 二聚体以上 的分析 TSKgel UltraSW Aggregate 1.mL/.2mL/ TSKgel UltraSW Aggregate (7.8 mm 3 cm ID..2 mol/l (ph 6.7 +.5 % NaN3.2 1. ml/ UV (28 nm 25 μl 5 g/l 4 6 8 3 4 ~ 4 ~
1-5 SEC 法对单链抗体 (scfv 的分析 G3SWXL scfv µg scfv + % (v/v x concentrated HCP solution A.1 M acetate +.2 M NaCl ph 3.5 B.1 M acetate +.5 M NaCl ph 5.6 45 CV 4 % 75 B CV % B Toyopearl MX-Trp-65M TSKgel G3SW XL.1 M sodium phosphate sodium +.1 M sulfate ph 6.7 1-6 SEC 法对各种抗体药物的分析实例 I TSKgel G3SWXL 2. 35 3 25 15 5 Rs 2 TSKgel G3SW XL (7.8 mm ID 3 cm ( mmol/l 2PO4 + KH mmol/lna2so4 ph 6.6:IPA=9:1 1. ml/ μl R.T. A ( 2 4 6 8 12 14 16 18 ~ 5 ~
1-7 SEC 法对各种抗体药物的分析实例 II TSKgel G3SWXL 2. 4 3 4 3 2 1 Rs 2 TSKgel G3SW XL (7.8 mm ID 3 cm 5 mmol/lpb+3 mmol/lnacl (ph 6.8.8 ml/ μl R.T. B ( -1 2 4 6 8 12 14 16 18 2 4 6 8 12 14 16 18 1-8 SEC 法对各种抗体药物的分析实例 III TSKgel G3SWXL 2. 4 3 4 3 2 1 Rs 2 TSKgel G3SW XL(7.8 mm ID 3 cm 5 mmol/lpb+3 mmol/lnacl (ph 6.8.8 ml/ μl -1 2 4 6 8 12 14 16 18 R.T. C ( 2 4 6 8 12 14 16 18 ~ 6 ~
1-9 SEC 法对各种抗体药物的分析实例 IV TSKgel G3SWXL 4 3 3 25 15 5-5 - 5. 5.5 6. 6.5 7. 7.5 8. 8.5 9. 9.5. TSKgel G3SW XL(7.8 mm ID 3 cm 5 mmol/lpb+3 mmol/lnacl (ph 7. 1. ml/ μl R.T. D ( 2 4 6 8 12 14 16 18 1- SEC 法对各种抗体药物的分析实例 V TSKgel SuperSW mab HR 2. 8 6 4 4 3 2 1 Rs 2 TSKgel SuperSW mab HR (7.8 mm ID 3 cm mmol/lkh 2PO4 + mmol/lna2so4 (ph 6.6 1. ml/ μl R.T. E ( 2 2 4 6 8 12 14 2 4 6 8 12 14 ~ 7 ~
TOSOH BIOSCIENCE 1-11 SEC 法对各种抗体药物的分析实例 VI TSKgel SuperSW mab HR 1.5 4 35 3 25 15 Norm. 12 8 6 4 2-2 Rs 1.5 2 4 6 8 12 14 16 18 TSKgel SuperSW mab HR (7.8 mm ID 3 cm mmol/lpb+ mmol/lnacl (ph 6.8.8 ml/ μl R.T. F ( 5 2 4 6 8 12 14 16 18 1-12 SEC SuperSW3 Half-Fab mab TSKgel SuperSW3 4 μm (4.6 mm ID 3 cm mmol/l+ mmol/l ph 6.7 +.5% NaN3.35 ml/ FLD (λex: 28 nm λem: 35 nm 3 μl ~ 8 ~
1-13 IEC 法对抗体的分析对比实例 SP-STAT CM-STAT TSKgel SP-STAT TSKgel CM-STAT (4.6 mm ID cm A mmol/l (ph 7. B mmol/l +5 mmol/l NaCl (ph 7. -% B (3 UV28 nm 1 ml/ μl G ( 1-14 IEC 法对抗体中电荷异构体的分析 I TSKgel SP-STAT 9 Norm. 14 TSKgel SP-STAT (4.6 mm ID cm 12 A mmol/lpb (ph 6.5 B mmol/lpb + 1 mol/l NaCl (ph 6.5 ( 8 5 25 3 3.1 35 6 B% 4.5 ml/ 2 μl 6 7 8 9 11 12 13 14 15 H ( ~ 9 ~
TOSOH BIOSCIENCE 1-15 IEC 法对抗体中电荷异构体的分析 II TSKgel CM-STAT 8 12 11 9 8 7 6 5 4 3 2 1-1 -2 2 4 6 8 12 14 16 18 22 24 26 28 3 TSKgel CM-STAT (4.6 mm ID cm A mmol/lhac-naac (ph 6. B mmol/lhac-naac + 1 mol/l NaCl (ph 6. ( 1 125 125.1 3.1 13 B% 25.5 ml/ μl I ( 1-16 IEC 法对抗体中电荷异构体的分析 III TSKgel CM-STAT 7 4 35 3 25 TSKgel CM-STAT (4.6 mm ID cm A mmol/lpb (ph 7.5 B mmol/lpb + mmol/l NaCl(pH 7.5 ( 5 43 44 46 46.1 55 15 5 3 4 5 B% 3.8 ml/ μl J ( ~ ~
1-17 IEC 法对抗体中电荷异构体的分析 IV TSKgel Q-STAT 6 Norm. 8 6 TSKgel Q-STAT (4.6 mm ID cm A mmol/ltris-hcl (ph 9. B mmol/ltris-hcl + 1 mol/l NaCl (ph 9. (.1 24 24.1 3 4 B% 8 55 5 5 15 25.5 ml/ μl K ( 1-18 HILIC 法分离单克隆抗体的糖链 (PA 化寡糖 Rs=1.26 TSKgel Amide-8 3 μm (2. mm ID. 15 cm A mmol/l (ph + 7.3 B 75%B 75 (-5 5% B (5-8.5 ml/ (Ex.315 nm Em.38 nm 4 5 μl PA ( 5 15 25 3 ~ 11 ~
1-19 2-AB 标记糖链的 LC-MS 分析 TSKgel Amide-8 2μm (2. mm I.D. 15 cm A 5 mmol/l (ph 5.7 B 75%B (-5 75-5%B (5-3.3 ml/ 4 2-AB N IgG ( 8 pg (8% (A ex. 33 nm 4 (B MS ESI positive SIM 2-1 SEC 法对抗体片段的分析实例 I TSKgel SuperSW mab HR 1.8 225 175 15 125 75 5 25 Rs 1.8 2 4 6 8 12 14 16 18 TSKgel SuperSW mab HR (7.8 mm ID 3 cm mmol/lpb+ mmol/lnacl (ph 6.8.8 ml/ μl R.T. A ( ~ 12 ~
2-2 SEC 法对抗体片段的分析 II TSKgel SuperSW mab HR 1.8( ACN 225 175 15 125 75 5 25 Rs 1.8 2 4 6 8 12 14 16 18 TSKgel SuperSW mab HR (7.8 mm ID 3 cm mmol/lpb+ mmol/lnacl (ph 6.8 : ACN= 9:1.8 ml/ μl R.T. B ( 2-3 SEC 法对抗体片段的分析 III TSKgel SuperSW mab HR 2. 7 6 5 4 3 TSKgel SuperSW mab HR (7.8 mm ID 3 cm mmol/lpb+ mmol/lnacl(ph 6.8:ACN=9:1.8 ml/ μl R.T. C ( Rs 2 2 4 6 8 12 14 16 18 ~ 13 ~
2-4 IEC 法对抗体片段中异构体的分析 I TSKgel CM-STAT 2. 7 TSKgel CM-STAT (4.6 mm cm ID 6 5 4 3 Rs 2 5 15 25 A 5 mmol/lpb (ph 5. B 5 mmol/lpb + 1 mol/l NaCl (ph 5. ( 5 25 25 25.1 3 B%.8 ml/ μl D ( 2-5 IEC 法对抗体片段中异构体的分析 II TSKgel Super Q-5PW 5 6 5 4 3 TSKgel Super Q-5PW (7.5 mm ID 7.5 cm A mmol/ltris-hcl (ph 8.5 B mmol/ltris-hcl + 1 mol/l NaCl (ph 8.5 ( 2 32 35 35.1 4 B% 5 5 5 5 15 25 3 35 4.6 ml/ μl E ( ~ 14 ~
2-6 HIC 法对抗体片段的分析实例 I 4 35 3 25 15 Antibody TSKgel Phenyl-5PW (5 mm ID 5 cm A mmol/ltris-hcl + 242SO4 mol/l (ph (NH8.6 B mmol/ltris-hcl (ph 8.6 ( 5 25 25.1 3 5 5 15 25 3 35 B%.6 ml/ μl F ( 15 Fragments 5 5 15 25 3 35 2-7 HIC 法对抗体片段的分析实例 II TSKgel Butyl-NPR 7 25 15 5 5 1 15 2 3 4 5 6 7 25 TSKgel Butyl-NPR (4.6 mm ID cm A mmol/ltris-hcl + 242SO4 mol/l (ph (NH8. B mmol/ltris-hcl (ph 8. ( 5 25.1 3 B% 3 3 75 3 3.5 ml/ μl G ( ~ 15 ~
2-8 HIC 法对抗体片段的分析实例 III TSKgel Butyl-NPR 15 6 5 4 3 3 2 1-1 -2-3 -4 5 15 25 5 15 25 TSKgel Butyl-NPR (4.6 mm ID cm A mmol/ltris-hcl + 242SO4 mol/l (ph (NH8. B mmol/ltris-hcl (ph 8. ( 5 25 25.1 3 B% 4 4 4 4.6 ml/ μl H ( 2-9 HIC 法对抗体片段的分析实例 IV TSKgel Butyl-NPR 2 1 8 6 TSKgel Butyl-NPR (4.6 mm ID cm A mmol/ltris-hcl + 242SO4 mol/l (ph (NH8.6 B mmol/ltris-hcl (ph 8.6 ( 5 25 25.1 4 B% 4 5 15 25 3 35.6 ml/ μl I ( ~ 16 ~
2- HIC 法对抗体片段的分析实例 V TSKgel Butyl-NPR 6 Norm. 15 5 5 15 25 TSKgel Butyl-NPR (4.6 mm ID cm A 25 mmol/lpb + 1.542SO4 mol/l (ph (NH7.3 B 25 mmol/lpb (ph 7.3 ( 3 32 32.1 4 B%.5 ml/ μl J ( 2-11 HIC 法对抗体片段的分析实例 VI TSKgel Butyl-NPR 7 6 5 4 TSKgel Butyl-NPR (4.6 mm ID cm A mmol/ltris-hcl + 242SO4 mol/l (ph (NH8. B mmol/ltris-hcl (ph 8. ( 5 25 25.1 3 3 B% 5 15 25.5 ml/ μl K ( ~ 17 ~
2-12 RPC 法对抗体片段的分析实例 I TSKgel Protein C4-3 11 25 15 5-5 - 5 15 25 TSKgel Protein C4-3 (4.6 mm 15 cm ID A H 2O.1%TFA B ACN.1%TFA ( 5 5.1 25.1 25.2 3 B% 25 4 45 UV214 nm 1. ml/ 55 μl L ( 25 25 2-13 RPC 法对抗体片段的分析实例 II Fab Fc F(ab 2 Fc Fab F(ab 2 TSKgel Protein C4-3 (4.6 mm 15 cm ID A H 2O/ACN/TFA = 9//.5 (v/v/v B H 2O/ACN/TFA = /8/.5 (v/v/v % - % B in 45 4 UV2 nm μl IgG 14 15 16 17 18 19 21 22 23 24 ~ 18 ~
2-14 RPC 法分离抗体 IgG 的重链和轻链 TSKgel ProteinC4-3 IgG-A IgG-B IgG-A IgG-B IgG-B ABS @ 215 nm 3 25 15 5 light chain light chain hydrophilic variant heavy chain heavy chain IgG-A IgG-B TSKgel Protein C4-3 ( 4.6 mm 15 cm ID A H 2O/ACN/TFA = 9//.5 (v/v/v B H 2O/ACN/TFA = /8/.5 (v/v/v % % B (45 5 UV215nm μl IgG-A ( IgG-B ( 14 15 16 17 18 19 21 22 23 24 ( 3-1 HIC 法分离曲妥珠单抗和 ADC 药物 Trastuzumab 5 15 25 3 DAR=2 DAR=4 ADC DAR= DAR=6 TSKgel Butyl-NPR (4.6 mm ID. cm A 25 mmol/l (ph 7. + 1.5 mol/l B 25 mmol/l (ph 7. / 2- = 8 / 2 % B.5 ml/ UV (28 nm μl.24 g/l ADC (Herceptin-vcMMAE 2.2 g/l DAR=8 5 15 25 3 ~ 19 ~
3-2 SEC 法分离 ADC 药物 SEC ADC UP-SW3 ADC. AU.18.16.14.12..8.6.4.2. -.2 UP-SW3 SuperSW mab HR G3SWXL..5 11. 11.5 12. 12.5 13. 13.5 14. 14.5 15. 15.5 16. 16.5 17. 17.5 18. 18.5 Min TSKgel UP-SW 3 (4.6 mm ID 3 cm TSKgel Super SW mab HR (4.6 mm ID 3 cm TSKgel G3SWXL (7.8 mm ID 3 cm (5 mmol/l PB + 5 mmol/l NaCl + mmol/l Arg.:IPA=95:5.35 ml/(up-sw3.5 ml/ (SuperSW mab HR.5 ml/ (G3SWXL 5 μl R.T ADC 1 1 mg/ml 3-3 有机溶剂对 ADC 在分离效果的影响 ADC % IPA 3 25 15%IPA 15 %IPA 5 25%IPA 5 15 25 3 TSKgel Butyl-NPR (4.6 mm ID cm A.25 mol/lpb+1.5 42SO4(pH mol/l (NH7. B.25 mol/lpb:ipa=different ratio (ph 7. %B ( UV28 nm.5 ml/ μl ADC 1 1.2 mg/ml ~ ~
3-4 ph 梯度 盐浓度对 ADC 分离效果的影响 TSKgel SP-STAT ADC 2 2 ph 11 14 12 8 6 4 2 12 8 6 4 2 5 5.5 6 6.5 7 7.5 8 8.5 9 9.5 8 6 4 2 5 15 25 5 15 25 3 35 TSKgel SP-STAT (4.6 mm ID cm A 25 mmol/lnaac-hac (ph 4.5 B A+1 mol/l NaCl (ph 4.5.5 ml/ μl R.T ADC 2.8 mg/ml TSKgel SP-STAT (4.6 mm ID cm A mmol/lmes+25 mmol/lhepes + mmol/l NaCl (ph 4.5 B mmol/lmes+25 mmol/lhepes + mmol/l NaCl (ph.8.5 ml/ μl R.T ADC 2.8 mg/ml ~ 21 ~
TOSOH BIOSCIENCE 产品一览表 ( TSKgel UP-SW3 TSKgel UP-SW3 TSKgel SuperSW mab HR TSKgel SuperSW mab HTP TSKgel UltraSW Aggregate TSKgel SuperSW TSKgel SuperSW3 TSKgel SuperSW3 TSKgel SuperSW3 TSKgel GSW XL TSKgel G3SW XL TSKgel G4SW XL 2 μm 2 μm 4 μm 4 μm 3 μm 4 μm 4 μm 4 μm 4 μm 5 μm 5 μm 8 μm 4.6 mm ID 15 cm 4.6 mm ID 3 cm 4.6 mm ID 15 cm 7.8 mm ID 3 cm 7.8 mm ID 3 cm 4.6 mm ID 3 cm 1. mm ID 3 cm 2. mm ID 3 cm 4.6 mm ID 3 cm 7.8 mm ID 3 cm 7.8 mm ID 3 cm 7.8 mm ID 3 cm 5 5 5 5 5 5 5 5 2 5 6 1 5 5 5 5 5 5 5 1 5 5 5 7 6 23449 23448 22854 22855 22856 18674 21845 21485 18675 8541 854 8542 TSKgel Q-STAT 7 μm 4.6 mm I.D.. cm 27μeq/g-drygel Cl - 21961 TSKgel Q-STAT TSKgel SP-STAT μm 7 μm 3. mm I.D. 3.5 cm 4.6 mm I.D.. cm 27μeq/g-drygel 23μeq/g-drygel Cl - Na + 2196 21964 TSKgel SP-STAT μm 3. mm I.D. 3.5 cm 23μeq/g-drygel Na + 21963 TSKgel CM-STAT 7 μm 4.6 mm I.D.. cm μeq/g-drygel Na + 21966 TSKgel CM-STAT μm 3. mm I.D. 3.5 cm μeq/g-drygel Na + 21965 TSKgel DEAE-NPR 2.5 μm 4.6 mm I.D. 3.5 cm.1 eq/l Cl - 1375 TSKgel DNA-NPR TSKgel SP-NPR TSKgel SP-NPR 2.5 μm 2.5 μm 2.5 μm 4.6 mm I.D. 7.5 cm 4.6 mm I.D. 3.5 cm 4.6 mm I.D. 5. cm.1 eq/l.1 eq/l.1 eq/l Cl - Na + Na + 18249 1376 21942 (nm TSKgel Phenyl-5PW TSKgel Ether-5PW TSKgel Butyl-NPR TSKgel Butyl-NPR μm μm 2.5 μm 2.5 μm 7.5 mm I.D. 7.5 cm 7573 7.5 mm I.D. 7.5 cm 8641 4.6 mm I.D. 3.5 cm 14947 4.6 mm I.D. cm 42168 ~ 22 ~
TSKgel Amide-8 TSKgel Amide-8 TSKgel Amide-8 TSKgel Amide-8 TSKgel Amide-8 TSKgel Amide-8 TSKgel Amide-8 TSKgel Amide-8 TSKgel Amide-8 TSKgel Amide-8 2 μm 2 μm 2 μm 2 μm 2 μm 2 μm 3 μm 3 μm 3 μm 3 μm 2. mm ID 5 cm 2. mm ID cm 2. mm ID 15 cm 3. mm ID 5 cm 3. mm ID cm 3. mm ID 15 cm 2. mm ID 5 cm 2. mm ID 15 cm 4.6 mm ID 5 cm 4.6 mm ID 15 cm 23454 23455 23456 23457 23458 23459 21864 21865 21866 21867 TSKgel Protein C4-3 TSKgel Protein C4-3 TSKgel Protein C4-3 TSKgel Protein C4-3 TSKgel Protein C4-3 TSKgel Protein C4-3 3 μm 3 μm 3 μm 3 μm 3 μm 3 μm 2. mm I.D. 5 cm 2. mm I.D. cm 2. mm I.D. 15 cm 4.6 mm I.D. 5 cm 4.6 mm I.D. cm 4.6 mm I.D. 15 cm 2283 22831 22832 22827 22828 22829 ~ 23 ~
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