03

Size Exclusion Columns

YMC-SEC MAB ---------------------------------------- 32-33
YMC-Pack Diol ---------------------------------------- 34-35
Ordering Information------------------------------------- 36

03

Size Exclusion Columns
Size exclusion chromatography column for separation of monoclonal antibody (MAb)

YMC-SEC MAB
l Silica-based SEC column optimized for separation of MAbl Suitable for separation of aggregates and fragments of MAbl Excellent resolution and peak shapes

Particle size : 3 μm
Pore size : 250 Å■ Usable pH range : 5-7.5■ USP L20, L59■■Size Exclusion Columns

Size exclusion chromatography column for separation of monoclonal antibody (MAb)
YMC-SEC MAB is a silica-based size exclusion chromatography (SEC)/gel filtration chromatography (GFC) column with excellent
durability and batch-to-batch reproducibility. Optimized base material design and surface modification provide superior separationof antibody monomers and their aggregates/fragments with very low nonspecific adsorption. This is ideal for the quality control
assessment of antibody drugs.

Specifications
YMC-SEC MAB
Base

Silica gel

Particle size (μm)

3Pore size (Å)

250Bonded phase

Dihydroxypropyl

Usable pH range

5-7.5

Molecular weight range

10,000-700,000

Suitable for separation of aggregates and fragments of MAb
Analysis of monoclonal antibody aggregates
mAU
monomermAU
1200

Column

1000

30800

20

aggregates

10600

0400

10121416200
0M180604AA

532: YMC-SEC MAB (3 μm, 250 Å)
300 X 4.6 mmI.D.
Eluent: 0.1 M KH2PO4-K2HPO4 (pH 7.0)
containing 0.2 M NaCl
Flow rate: 0.165 mL/min
Temperature : 25°CDetection: UV at 280 nm
Injection: 10 μLSample: Humanized monoclonal antibody (5 mg/mL)

40

10

15

20

25min

182022 min

Humanized monoclonal antibody was analyzed using
YMC-SEC MAB. The chromatogram shows excellent
peak shape, and good separation between aggregates
and a monomer peak. YMC-SEC MAB is effective
for characterization and quality control of antibody
therapeutics.

Analysis of monoclonal antibody fragments
mAU

Intact monoclonal antibody
Digested monoclonal antibody

1175150

2. Fab dimer

3. Fc

4. Fab*1254100

1. Intact antibodyColumn

50: YMC-SEC MAB (3 μm, 250 Å)
300 X 4.6 mmI.D.
Eluent: 0.1 M KH2PO4-K2HPO4 (pH 7.0)
containing 0.2 M NaCl
Flow rate: 0.165 mL/min
Temperature : 25°C
Detection: UV at 280 nm
Injection: 2 μL (3 mg/mL)

3252M180802EA0M180802EA701012141618202224min*Peak derived from papain digestive fluid

A monoclonal antibody digested by papain and an intact monoclonal antibody were analyzed using YMC-SEC MAB. Since the column shows excellent
separation between Fc and Fab fragments, which have similar amount of MW, it is effective for separation of antibody fragments.

Size Exclusion Columns

75

Batch-to-batch reproducibilityColumn: YMC-SEC MAB (3 μm, 250 Å)
300 X 4.6 mmI.D.
Eluent: 0.1 M KH2PO4-K2HPO4 (pH 7.0)
containing 0.2 M NaClFlow rate
: 0.165 mL/min
Temperature : 25°CDetection: UV at 280 nm
Injection: 10 μL
Sample: Humanized monoclonal antibody (5 mg/mL)

mAU

40M180604BA2Batch A

M180525AA10Batch B

3020YMC-SEC MAB provides excellent reproducibility of theseparation of monomer and aggregates as well as formonomer and fragments. Therefore, it is very effective for
quality control of antibody drugs.

10Batch C

M180604AA

010121416182022min

Excellent durability

M180717AH50

20400thM180717AF50151050100%

8.595%

890%

7.50M180717AD50200th

M180717AB50100th
1st246810min

100200300

40085%500Run number

300thM180717AA010-5500th

9% of initial theoreticalplate number

M180717AJ5025105%

Elution time (min)mAU

9.5Column: YMC-SEC MAB (3 μm, 250 Å)
300 X 4.6 mmI.D.
Eluent: 0.1 M NaH2PO4-Na2HPO4 (pH 7.0)
containing 0.2 M NaCl
Flow rate: 0.33 mL/min
Temperature : 25°C
Detection: UV at 280 nm
Injection: 2 μL
Sample: Humanized monoclonal antibody (1 mg/mL)

YMC-SEC MAB has excellent repeatability of separation without any change of theoretical plate number and elution time even after repeated separationof monoclonal antibody for more than 500 times because of its superior durability.

33

03

Size Exclusion Columns
Silica-based size exclusion chromatography columns

YMC-Pack Diol‌Particle size : 2, 3, 5 μm
‌Pore size : 60, 120, 200, 300 Å‌Usable pH range : 5-7.5
‌USP L20, L33, L59

 ‌Silica-based columns with high mechanical stability ‌Useful for molecular weight determination of proteins and sugars ‌High-resolution and fast analysis using 2 μm, 3 μm particle

Size Exclusion Columns

Silica-based size exclusion chromatography (SEC) column
YMC-Pack Diol is a size exclusion chromatography column packed with dihydroxypropyl-bonded silica, and available in four different
pore sizes. Diol-120, 200, and 300 are suitable for separation or molecular weight determination of proteins with molecular weights of
1,000 to 1,000,000. Diol-60 is the most suitable for separation of peptides or oligosaccharides whose molecular weights are 10,000 orless.

SpecificationsColumnBase

Bonded phase

Pore size(Å)

Diol-60

Particle size(μm)

UsablepH range60

Characteristics
For molecular weights less than 10,000

3, 5Diol-120

120
Silica gel

For molecular weights 1,000 to 100,000

Dihydroxypropyl

Diol-2005-7.5200

For molecular weights 5,000 to 300,000
2, 3, 5Diol-300

300

For molecular weights 20,000 to 1,000,000

Calibration curves of various proteins
Comparison of different pore sizes (5 μm)
106Comparison of different particle sizes (Diol-300)
106

125 μm

343 μm2 μm

105
Diol-120

Diol-200

Diol-300

MWMW56105

78910111214104
57Column

1315169
Elution volume (mL)

11: YMC-Pack Diol
300 X 8.0 mmI.D.
Eluent: 0.1 M KH2PO4-K2HPO4 (pH 7.0)
containing 0.2 M NaCl
Flow rate: 0.5 mL/min
Temperature : 25°C
Detection: UV at 280 nm

13104

2.0

2.5

3.0
3.5
Elution volume (mL)

4.0

MW
1. IgM
900,000
2. Thyroglobulin
670,000
3. IgA
390,000
4. Fibrinogen
340,000
5.γ-Globulin
158,000
6. IgG
150,000
7. Transferrin
75,0008. HSA (human serum albumin) 66,000
9.α1-Antitrypsin
50,00010. Ovalbumin
45,00011. Carbonic anhydrase
30,00012. Trypsin inhibitor
20,10013. Myoglobin
17,00014.α-Lactalbumin
14,10015. Ribonuclease A
13,70016. Cytochrome c
12,400Column: YMC-Pack Diol
300 X 4.6 mmI.D.
Eluent: 0.1 M KH2PO4-K2HPO4 (pH 7.0)
containing 0.2 M NaClFlow rate
: 0.165 mL/min
Temperature : 25°CDetection: UV at 280 nm

Diol-120, Diol-200, and Diol-300 are suitable for the separation or molecular weight determination of proteins with molecular weights from 1,000 to 1,000,000.
Calibration curves of proteins with different particle sizes using YMC-Pack Diol are almost the same and this enables easy method transfer between
UHPLC and HPLC.

34

Separation for standard protein markers
Diol-120

Diol-200

Diol-300

141 3524512310420030 min

10Column: YMC-Pack Diol
500 X 8.0 mmI.D.
Eluent: 0.1 M KH2PO4-K2HPO4 (pH 7.0)
containing 0.2 M NaClFlow rate
: 0.7 mL/min
Temperature : ambient
Detection: UV at 280 nm

52030040min102030

For molecular weight 5,000 to 300,000 substances,
Diol-200 is suitable for the separation.

40 minSize Exclusion Columns

230MW
290,000
142,000
67,00032,00012,4001. Glutamate dehydrogenase
2. Lactate dehydrogenase
3. Enolase
4. Adenylate kinase
5. Cytochrome c

Separation of peptides with molecular weights less than 10,000
Diol-60
123

MW
5,700
1,672
1,04075

1. Insulin (Bovine)
2. Neurotensin
3. Angiotensin II
4. Glycine

412104345Column

010203040 min: YMC-Pack Diol-60
500 X 8.0 mmI.D.
Eluent: 0.1 M KH2PO4-K2HPO4(pH 7.0)
containing 0.2 M NaCl/acetonitrile (70/30)
Flow rate: 0.7 mL/min
Temperature : ambientDetection: UV at 215 nm

79103
MW

1214156811131016102

1710

For molecular weight less than 10,000 peptides, Diol-60 is suitable for the separation.

15Elution volume (mL)MW
1. Myoglobin
17,0002. Ribonuclease A
13,7003. Cytochrome c
12,4004. Insulin (Bovine)
5,700
5. Insulin B chain
3,496
6. α-Mating factor
1,684
7. Neurotensin
1,672
8. Angiotensin I
1,296
9. CCK-Octapeptide
1,143
10. Bradykinin
1,060
11. Angiotensin II
1,040
12. Oxytocin
1,007
13. Angiotensin III
931
14. Met-Enkephalin
574
15. Leu-Enkephalin
555
16. Tetraglycine
246
17. Glycine
75

20

Separation of oligo- and polysaccharides
Diol-200
1MW
1. P-800 853,000
2. P-50
48,000
3. P-20
23,700
4. P-5
5,800

105

1
Diol-300

234
62

104

Diol-20

7

9
Diol-60102

101020 minColumn: YMC-Pack Diol, 500 X 8.0 mmI.D.
Eluent: water
Flow rate: 1.0 mL/min
Temperature : ambient
Detection: RI081030MW
1. Pullulan (P-800)
853,000
2. Pullulan (P-400)
380,000
3. Pullulan (P-200)
186,000
4. Pullulan (P-100)
100,000
5. Pullulan (P-50)
48,000
6. Pullulan (P-20)
23,700
7. Pullulan (P-10)
12,2008. Pullulan (P-5)
5,800
9. Maltopentadecaose (G15) 2,448
10. Maltoundecaose (G11)1,800
11. Maltoheptaose (G7)1,152
12. Maltopentaose (G5)824
13. Maltotriose (G3)504
14. Maltose (G2)342
15. Glucose (G1)
180

54MW

3

10610Diol-1

11121320141515
Elution volume (mL)2025

For separation or molecular weight determination of water-soluble oligo- and polysaccharides, Diol-60,Diol-120, Diol-200, and Diol-300 are useful individually or in combination.

35

03

Size Exclusion Columns
Ordering Information –Columns–
YMC-SEC MAB
Particle size
(μm)

Column size
inner diameter X length (mm)YMC-SEC MAB(250 Å)4.6 X 150

DLM25S03-1546WT

4.6 X 300

DLM25S03-3046WT

8.0 X 300

DLM25S03-3008WT3

Size Exclusion Columns

YMC-Pack Diol (Stainless steel columns)
Particlesize(μm)
235Column size
inner diameter X length (mm)Diol-60(60 Å)

Diol-120(120 Å)

Diol-200(200 Å)

Diol-300(300 Å)4.6 X 150——

DL20S02-1546PTH

DL30S02-1546PTH

4.6 X 300——

DL20S02-3046PTH

DL30S02-3046PTH

4.6 X 300

DL06S03-3046WTDL12S03-3046WTDL20S03-3046WT

DL30S03-3046WT

4.6 X 300

DL06S05-3046WTDL12S05-3046WTDL20S05-3046WT

DL30S05-3046WT

8.0 X 300

DL06S05-3008WTDL12S05-3008WTDL20S05-3008WT

DL30S05-3008WT

8.0 X 500

DL06S05-5008WTDL12S05-5008WTDL20S05-5008WT

DL30S05-5008WT

20 X 300

DL06S05-3020WT

DL12S05-3020WT

DL20S05-3020WT

DL30S05-3020WT

20 X 500

DL06S05-5020WT

DL12S05-5020WT

DL20S05-5020WT

DL30S05-5020WT

YMC-Pack Diol Guard columns (Stainless steel columns)
Particlesize(μm)
5Column size
inner diameter X length (mm)Diol-60(60 Å)

Diol-120(120 Å)

Diol-200(200 Å)

Diol-300(300 Å)

8.0 X 30

DL06S05-0308WTG

DL12S05-0308WTG

DL20S05-0308WTG

DL30S05-0308WTG

20 X 50

DL06S05-0520WTG

DL12S05-0520WTG

DL20S05-0520WTG

DL30S05-0520WTG

Column size
inner diameter X length (mm)Diol-60(60 Å)

Diol-120(120 Å)

Diol-200(200 Å)

Diol-300(300 Å)

8.0 X 300

DL06S05-3008FG

DL12S05-3008FG

DL20S05-3008FG

DL30S05-3008FG

8.0 X 500

DL06S05-5008FG

DL12S05-5008FG

DL20S05-5008FG

DL30S05-5008FG

YMC-Pack Diol (Glass columns)
Particlesize(μm)
536