MonoCap HighResolution 2000 Series
Optimized for Identification of Peptides /Proteinsfor Proteome Research
MonoCap HighResolution 2000 is a 2 meter length monolithic silica capillary column which is designed for identifying extremely high number of peptides/proteins for proteome research via LC-MS/MS.
What is MonoCap HighResolution 2000 ?
MonoCap C18 HighResolution 2000Silica : High Purity Monolithic Silica GelThrough-pore Size : 2 μmMesopore Size : 15 nmBonded Phase : Octadecyl GroupsEnd-capping : YesInternal Diameter : 0.1 mm I.D.Length : 2000 mmRecommended Flow Rate Ranges : 0.3 μL/min ~ 1 μL/minMaximum Operating Pressure : 35 MPa, 300 Bar
Physical Properties of MonoCap HighResolution 2000
MonoCap HILIC-UP HighResolution 2000Silica : High Purity Monolithic Silica GelThrough-pore Size : 2 μmMesopore Size : 12 nmBonded Phase : Ureidopropyl GroupsEnd-capping : NoneInternal Diameter : 0.1 mm I.D.Length : 2000 mmRecommended Flow Rate Ranges : 0.3 μL/min ~ 1 μL/minMaximum Operating Pressure : 35 MPa, 300 BarRemarks : For identification of highly hydrohophilic peptides/proteins
Mesopores Through-pores
Bimodal Pore Structure of MonoCap
Why Use MonoCap HighResolution 2000 columns?
Facts
Separation Efficiency Higher Lower
Number of Identified Peptides More Less
Total Operation Time Shorter Longer
Operating Pressure Lower Higher
Column Lifetime Longer Shorter
Total Cost including all the Less Moreabove factors
MonoCap HighResolution2000 mm columns
Traditional Particle Packed150 mm columns
Performance of MonoCap C18 HighResolution 2000
Column NameNumber of
Identified Proteins in average
Analysis Time
MonoCap C18 HighResolution 20000.1 mm I.D. x 2000 mm
2,087 (2013, 2116, 2131) 10 Hours
Particle packed column0.1 mm I.D. x 150 mm
680(685, 679, 675) 2 Hours
MS: LTQ-Orbitrap XL (Mascot Search)Sample: THP-1 Cell Lysate Tryptic Digest, 5 μg
All data were acquired, provided by our customer
Performance of MonoCap C18 HighResolution 2000
180 200 220 240 260 280 300 320 340 360 380 400 420 440 460 480 500 520 540 560 580 600160140120100806040200
[1] MonoCap C18 HighResolution 2000 (2000 mm x 0.1 mm I.D.)
Time (min)
[2] Particle Packed column (3 μm , 150 mm x 0.075 mm I.D.)
Time (min)
ConditionsSystem Column
Trap column Eluent
: GLS Capillary HPLC system: [1] MonoCap C18 High Resolution 2000 (2000 mm x 0.1 mm I.D): [2] Particle packed column (3 μm, 150 mm x 0.075 mm I.D.): MonoCap C18 Trap Column (50 mm x 0.075 mm I.D.): A) 0.1 %HCOOH in CH3CN
B) 0.1 %HCOOH in H2O[1] A / B = 10 / 90 - 600 min - 45 / 55 [2] A / B = 10 / 90 - 180 min - 45 / 55
: [1] 0.5 μL/min[2] 0.3 μL/min
: 5 μL : MS (TIC m / z 500 - 1500): Tryptic digest of proteins
Flow Rate
Injection Vol.DetectionSample
● ● ● are analytes having the same molecular weight
Performance of MonoCap C18 HighResolution 2000[1] MonoCap C18 HighResolution 2000 (2000 mm x 0.1 mm I.D.)
Time (min)
[2] Particle Packed column (3 μm , 150 mm x 0.075 mm I.D.)
Time (min)
● ● ● are analytes having the same molecular weightConditionsSystem Column
Trap column Eluent
: GLS Capillary HPLC system: [1] MonoCap C18 High Resolution 2000 (2000 mm x 0.1 mm I.D): [2] Particle packed column (3 μm, 150 mm x 0.075 mm I.D.): MonoCap C18 Trap Column (50 mm × 0.075 mm I.D.): A) 0.1 %HCOOH in CH3CN
B) 0.1 %HCOOH in H2O[1] A / B = 10 / 90 - 600 min - 45 / 55 [2] A / B = 10 / 90 - 180 min - 45 / 55
: [1] 0.5 μL/min[2] 0.3 μL/min
: 5 μL : MS (TIC m / z 500 - 1500): Tryptic digest of proteins
Flow Rate
Injection Vol.DetectionSample
Analysis Time: 3 hrs
180 200 220 240 260 280 300 320 340 360 380 400 420 440 460 480 500 520 540 560 580 600160140120100806040200
180 200 220 240 260 280 300 320 340 360160140120100806040200
Performance of MonoCap C18 HighResolution 2000
Time (min)
Analysis Time: 6 hrs
Time (min)
Analysis Time: 10 hrs
Time (min)
● ● ● are analytes having the same molecular weightColumn : MonoCap C18 High Resolution 2000 (2000 mm x 0.1 mm I.D)Trap column : MonoCap C18 Trap Column (50 mm x 0.075 mm I.D.)Flow Rate : 0.5 μL/minEluent : A) CH3CN (0.1%HCOOH), B) H2O (0.1%HCOOH)
A / B = 10 / 90 - 180, 300, 600 min- 45/55Sample : Tryptic digest of proteinsInjection vol. : 5 μL
Performance of MonoCap C18 HighResolution 2000
Shotgun Proteomics study using MonoCap C18 HighResolution 2000 mm
Sample: Proteins derived from rat serum, brain microsome and T Cell
5 μL Loop Injection
[LTQ-XL]m/z 400-1500 top5 CID
[LTQ-Orbtirap XL]m/z 400-1500 top3 CID
[mascot search] DB: Refseq_rat, 16776 sequence (including only NP_******) LTQ-XL: cut off =35 ptsLTQ-OrbitrapXL: cut off=0.05
All data were acquired, provided by Astellas Pharma Inc., Mr. Hiramoto
Performance of MonoCap C18 HighResolution 2000
MonoCap C18 12hr gradient
LTQ-XL(Top5 CID)
MonoCap C18 6hr gradient
LTQ-XL(Top5 CID)
15 cm Particle packed2hr gradient
LTQ-OrbitrapXL(Top3 CID)
Offgel fractionator(12 frs)
15cm packed 2hrOrbitrapXL(Top3 CID)
Rat brain microsomeTryptic digestca.25 μg
1,772 1,661 915 2,800
Rat brain microsomeTryptic digestiTRAQ5 μg
1,487 1,182 210(Top3 HCDCID) -
Rat serum 0.4 μL(ca. 20 μg protein)Tryptic digest 329 232 153 -
Particle Packed 15 cm vs MonoCap C18 HighResolution 2000Comparison of Number of Identified Proteins
Performance of MonoCap C18 HighResolution 2000
RT: 0.00 - 119.98
0 10 20 30 40 50 60 70 80 90 100 110Time (min)
0
20
40
60
80
100
Rel
ative
Abu
ndan
ce
60.80621.6653
68.17588.3217
98.59416.3369
72.60644.3710 99.68
444.368076.91
923.506854.15
535.309796.48
581.3677
91.58816.4934
45.97612.2755
77.35599.3589
100.85415.211641.66
560.778031.53451.7808
84.38951.8036
12.76437.8893
105.10639.6283
0.18445.1200
18.59445.1200
NL:1.75E8Base Peak MS FMBM06_1H19_01
RT: 0.00 - 720.02
0 50 100 150 200 250 300 350 400 450 500 550 600 650 700Time (min)
0
10
20
30
40
50
60
70
80
90
100
Rel
ativ
e Ab
unda
nce
387.38766.51
367.42705.56
451.80852.40420.82
675.53350.08567.38
271.54622.08
191.38535.58
255.57579.37
638.10403.16501.49
817.06330.59588.57237.52
709.52 288.92652.57
463.58952.06
185.11598.18
505.79746.26
314.57607.68
485.90713.60
617.25430.34
137.24612.59
171.10470.58
130.21516.25
597.39416.55
220.07553.71
76.46452.14
29.49438.42 524.14
848.27704.42445.11
690.20445.13
NL:7.59E6Base Peak M S monollith2m_110902_06
RT: 0.00 - 360.01
0 20 40 60 80 100 120 140 160 180 200 220 240 260 280 300 320 340 360Time (min)
0
10
20
30
40
50
60
70
80
90
100
Rel
ativ
e Ab
unda
nce
152.34567.35
165.79766.50 178.50
924.20
189.57956.83
279.04822.64128.20
622.13150.51
588.63138.27652.57 212.24
817.04124.67579.51 285.89
563.40194.99952.12103.86
535.55 116.18709.54 224.15
581.54 259.91416.51
83.75612.54
74.82561.05
61.55452.09
250.42449.96
50.53471.05
287.57656.82
235.22515.97
40.16455.47
335.36445.02
6.15444.96
354.51444.94
319.68445.02
NL:1.26E7Base Peak M S monollith2m_110901_04
0.1 x 150 mm 3 μm particle packed column210 protein hit (OrbitrapXL, HCDCID top3)
Rat brain microsome 5 μg tryptic digest iTRAQ
0.1 x 2000 mm MonoCap C18 40 ℃ 6hr gradient1,187 protein hit(LTQ-XL CID top3)
0.1 x 2000 mm MonoCap C18 40 ℃ 12 hr gradient1,482 protein hit(LTQ-XL CID top3)
Performance of MonoCap C18 HighResolution 2000
Example of 2-Dimensional LC for Phosphoproteomics
Rat T Cell (1x10e8) Tryptic digest from protein
0.1 x 2000 mm MonoCap C18 NanoLC/MS/MS(24hr gradient、LTQ-OrbitapXLInjecting two times separately)
Purificatin of phosphopeptides via HAMMOC Method(Phosphoric acid, Ammonia, Pyrrolidine gradual elution)
Anti P-Tyr Ab
ElutePass through
0.1 × 150 mm silicaC18 3μm NanoLC/MS/MS(2hr gradient、LTQ-OrbitapXL)
SCX column(2 x 150mm)
0.1 × 150 mm Silica C18 3 μm NanoLC/MS/MS
(2hr gradient、LTQ-OrbitapXL)
96 fr.
Identified Phosphopeptides: 11,380 Hours of Operation: 48 hrs
267phosphopeptides
2D LC
MonoCap 2m
1/21/2
More identified phosphopeptides in less time compared to 2D LC.
GL Sciences’Phos-TiO Kit
Identified Phosphopeptides: 8,324 Hours of Operation: 200 hrs
For Identification of Highly Hydrophilic Peptides /Proteins
Performance of MonoCap HILIC-UP HighResolution 2000
6,500 5,5004,000
HILIC-UP C18
Column Size : 2000 mm x 0.1 mm I.D. Mass spectrometer : Triple TOF 5600Gradient : Two Gradient Programs, 4 hrs & 8 hrs
Reference:Hydrophilic Interaction Chromatography Using a Meter-Scale Monolithic silica capillary Column for Proteomics LC-MS, K Horie et al. Anal. Chem. 2014, 86, 3817-3824
Performance of MonoCap HILIC-UP HighResolution 2000
ConditionsColumn : MonoCap HILIC-UP High Resolution 2000Eluent : A) CH3CN : H2O=10/90 (0.5% CH3COOH)
B) CH3CN : H2O=95/5 (0.5% CH3COOH)A/B=0/100-(240 min)-20/80-(10 min)-100/0-(10 min)-100/0
Flow Rate : 0.5 μLInjection Vol. : 1 μL (1 mg/mL)Detection : TIC MS (m/z 300-1500)Sample : Tryptic Digest of Hela Cell Lysate, 5 ug
Number of Identified Peptides : 7,194 (14736 PSM*)Number of Identified Proteins : 2,201*Peptide Spectrum Match
Total Ion Chromatogram Base Peak Chromatogram
0
20000000
40000000
60000000
80000000
100000000
120000000
0 50 100 150 200 2500
500000
1000000
1500000
2000000
2500000
3000000
3500000
4000000
0 50 100 150 200 250
Time (min)
Inte
nsity
Inte
nsity
Reference:Results of MonoCap C18 HighResolution 2000
Number of Identified Peptides: 8,358Number of Identified Proteins: 1,992Gradient Program: 4 hrs
Time (min)
Ordering Information
Description Model Remarks Cat.No.
Zero Dead Volume Union U-435 For 1/16“ SUS Tubing 6010-72352
Column ConnectionPurchase the following connection kit when directly connecting the column to the injector of a nano-LC instrument.
Description Contents Cat.No.
MonoCap C18 HighResolution 2000 Connection Kit 1/16“ PEEK Ferrule, SUS Nut, Sleeve, 2 pcs each.1/32“ PEEK Ferrule, SUS Nut, Sleeve, 2 pcs each. 5020-10017
MonoCap C18 HighResolution 2000 Connection Kit
1/16“ PEEK Ferrule 1/32“ PEEK Ferrule1/16“ PEEK Sleeve 1/32“ PEEK Sleeve
1/32“ SUS Nut1/16“ SUS Nut
Installing MonoCap to the appropriate End-fittings1. Prepare either 1/16” or 1/32” end-fittings and sleeve included in the MonoCap
connection kit (Cat# 5020-10017).
Ferrule
Nut
PEEK Sleeve
Installing MonoCap to the appropriate End-fittings2. Insert the sleeve into a nut and ferrule. Then insert a fused silica capillary tube into
the sleeve.
Ferrule
Nut
PEEK Sleeve
Fused Silica Capillary Tube(To be prepared by thecustomer)
Installing MonoCap to the appropriate End-fittings3. Insert the pre-assembled fitting into the injection port. Then tighten the nut by two
or three turns using a wrench to make certain that the ferrule grabs the sleeve.
Wrench
Nut
PEEK Sleeve
Fused Silica Capillary Tube(To be prepared by thecustomer)
Installing MonoCap to the appropriate End-fittings4. Now the ferrule is fixed to the sleeve. Remove the fused silica capillary tube from the
sleeve.
The ferrule is now fixed to the sleeve
Nut PEEK Sleeve
Fused Silica Capillary Tube(To be prepared by thecustomer)
Installing MonoCap to the appropriate End-fittings5. Insert the MonoCap C18 HighResolution 2000 column into the sleeve. Then push the
sleeve all the way into the port for a zero dead volume connection that the assembly seats firmly. Finally, keep pushing the MonCap C18 column into the port direction and tighten the nut by two or three turns.
Nut
PEEK Sleeve
MonoCap C18HighResolution 2000
Installing MonoCap to the appropriate End-fittings6. Gently pull the MonoCap C18 column to the direction shown below in blue. Make
sure the column is not loose and it is firmly installed.
MonoCap C18HighResolution 2000
Column ConnectionPurchase the following union when using a tubing that generates any indirect connection to the column.
Description Model Remarks Cat.No.
Zero Dead Volume Union U-435 For 1/16“ SUS Tubing 6010-72352
Example of Column Connection using nanoViper
Pump
Eluent
Injector MonoCap C18HighResolution 2000 LC-MS/MS
UnionnanoViper
Installing MonoCap to nanoViper1. Prepare 1/16” end-fittings and sleeve included in the MonoCap connection kit
(Cat# 5020-10017).
Ferrule
Nut
PEEK Sleeve
2. Insert the sleeve into a nut and ferrule. Then insert a fused silica capillary tube into the sleeve.
Ferrule
Nut
PEEK Sleeve
Fused Silica Capillary Tube(To be prepared by thecustomer)
Installing MonoCap to nanoViper
3. Insert the pre-assembled fitting into the injection port. Then tighten the nut by two or three turns using a wrench to make certain that the ferrule grabs the sleeve.
Wrench
Nut
PEEK Sleeve
Fused Silica Capillary Tube(To be prepared by thecustomer)
Installing MonoCap to nanoViper
4. Now the ferrule is fixed to the sleeve. Remove the fused silica capillary tube from the sleeve.
The ferrule is now fixed to the sleeve
Nut PEEK Sleeve
Fused Silica Capillary Tube(To be prepared by thecustomer)
Installing MonoCap to nanoViper
UnionnanoViper
5. Connect the MonoCap C18 HighResolution 2000 column into union. Push the sleeve all the way into the union for a zero dead volume connection that the assembly seats firmly. Keep pushing the MonCap C18 column into the union and tighten the nut by two or three turns. Gently pull the MonoCap C18 to confirm if the column is not loose and it is firmly installed. Finally, fingertight the nanoViper to the injection port.
PEEK Sleeve
MonoCap C18HighResolution 2000
Installing MonoCap to nanoViper