Table 6.
CE-based analysis of protein glycosylation
| Analyte | Mode | Capillary | BGE | Detection | Label | Notes | Ref. |
|---|---|---|---|---|---|---|---|
| N-glycans of mAb1 | CGE | 48 capillary array, 50 cm each |
Applied Biosystems Pop-7™ |
LIF ex. 473 nm / em. 520 nm |
APTS | High-throughput glycan analysis | [112] |
| (1) Neu5Gc and (2) α1,3-Gal ontaining N-glycans |
Partial-filling CE | 40 (30) cm, 50 µm id DB-1 capillary |
100 mM Tris-acetic acid, 0.05% HPC, pH 7.0 |
LIF ex. 488 nm / em. 520 nm |
APTS | Reaction with (1) anti-Neu5Gc or (2) α-galactosidase inject prior to sample |
[115] |
| Non-human -glycans |
CGE | 40 (30) cm, 100 µm id DB-1 capillary |
100 mM Tris-borate, 5% PEG, pH 8.3 |
LIF ex. 325 nm / em. 405 nm |
2-AA | Analysis of commercially available mAbs |
[116] |
| α1,3-Gal containing N-glycans |
CGE | 60 (50) cm, 50 µm id eCAP NCHO coated |
Beckman Coulter Carbohydrate Separation Gel Buffer-N |
LIF ex. 488 nm / em. 520 nm |
APTS and AMAC |
Ultrasensitive detection method | [117] |
| N-glycans of mAb | CGE | 50 (40) cm, 50 µm id PVA coated |
Beckman Coulter Carbohydrate Separation Buffer Or 40 mM EACA-acetate, 0.2% HPMC |
LIF ex. 488 nm / em. 520 nm |
APTS | Glycans of mAbs from NS0 cells | [118] |
| N-glycans of fusion protein |
CE | 90, 60, or 43 cm | 0.7 M ammonia and 0.1 M EACA in 70% MeOH |
TOF-MS | APTS | Alkaline CE-MS method | [119] |
| N-glycans | (1) CGE (2) CE |
Various capillary lengths and coatings |
(1) Beckman glycan separation buffer or POP-7 polymer (2) 40 mM EACA, 131 mM acetic acid, pH 4 |
(1) LIF ex. 488 nm / em. 512 nm (2) TOF-MS |
APTS | CGE-LIF and CE-MS methods compared to the CE-MS method in ref [119] |
[120] |
| N-glycans of mAbs | CE | 50 (40) cm, 50 µm id N-CHO coated |
Beckman Coulter Carbohydrate Separation Gel Buffer |
LIF ex. 488 nm / em. 520 nm |
APTS | CE-LIF as an orthogonal technique to MS |
[121] |
Capillary: Actual length (effective length), inner diameter, coating
2-aminoacridone (AMAC), 2-aminobenzoic acid (2-AA), 9-aminopyrene-1,3,6-trisulfonic acid (APTS), ε-aminocaproic acid (EACA), Galα1–3Gal (α1,3-Gal), Hydroxypropyl cellulose (HPC), Hydroxypropylmethylcellulose (HPMC), N-Glycolylneuraminic acid (Neu5Gc), Polyvinyl alcohol (PVA), Poly(ethylene glycol) (PEG),