![©2015 Waters Corporation 2
250 µg/mL Plasma
Time
1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 11.00
%
0
100
1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 11.00
%
0
100
1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 11.00
%
0
100
1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 11.00
%
0
100
04Apr2016_CRP_Cone_002 1: MRM of 3 Channels ES+
354.935 > 244.3 (AFVFPK)
8.62e6
Area
4.22;301179
04Apr2016_CRP_Cone_002 2: MRM of 4 Channels ES+
565.101 > 347.01 (ESDTSYVSLK)
1.35e7
Area
3.41;477497
04Apr2016_CRP_Cone_002 3: MRM of 2 Channels ES+
569.135 > 716.9 (GYSIFSYATK)
3.60e6
Area
4.60;119972
04Apr2016_CRP_Cone_002 4: MRM of 3 Channels ES+
697.287 > 567.1 (QDNEILIFWSK)
4.31e5
Area
5.84
9641
AFVFPK
ESDTSYVSLK
GYSIFSYATK
ACQUITY HSS T3 1.7 µm 2.1 x 50 mm
Peptide
Precursor Charge
State
MRM transition
Cone
Voltage
(V)
Collision
Energy
(eV)
Product Ion Identification
AFVFPK [M+2H]2+ 354.935>244.56 30 15 [2H+]1/y4 and [1H+]1/y2
ESDTSYVSLK [M+2H]2+ 565.101>347.011 15 15 [1H+]1/y3
GYSIFSYATK [M+2H]2+ 569.135>221.58 25 15 [1H+]1/b2
Quantification of Mid-Size Proteins:
CRP Tryptic Peptide LC-MS
For research use only. Not for use in diagnostic procedures](data:image/gif;base64,R0lGODlhAQABAIAAAAAAAP///yH5BAEAAAAALAAAAAABAAEAAAIBRAA7)
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Semelhante a Xevo TQ-XS Data: Quantifying Mid-size Proteins
Semelhante a Xevo TQ-XS Data: Quantifying Mid-size Proteins (20)
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Xevo TQ-XS Data: Quantifying Mid-size Proteins
- 1. ©2015 Waters Corporation 1 Quantification of Mid-Size Proteins: Human C-reactive Protein (CRP) CRP, MW ~25kDa, is an important clinical marker for inflammation, atherosclerosis, and infection Used as prognostic biomarker for cardiovascular risk and colorectal cancer 35 µL sample 3 Step Digestion (2 hours) Peptide purification with µElution SPE Clean Up Kit Human CRP Sequence MEKLLCFLVLTSLSHAFGQTDMSRKAFVFPK|ESDTSYVSLKAPLTKP LKAFTVCLHFYTELSSTRGYSIFSYATKRQDNEILIFWSKDIGYSFT VGGSEILFEVPEVTVAPVHICTSWESASGIVEFWVDGKPRVRKSLKKG YTVGAEASIILGQEQDSFGGNFEGSQSLVGDIGNVNMWDFVLSPDEI NTIYLGGPFSPNVLNWRALKYEVQGEVFTKPQLWP For research use only. Not for use in diagnostic procedures
- 2. ©2015 Waters Corporation 2 250 µg/mL Plasma Time 1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 11.00 % 0 100 1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 11.00 % 0 100 1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 11.00 % 0 100 1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00 11.00 % 0 100 04Apr2016_CRP_Cone_002 1: MRM of 3 Channels ES+ 354.935 > 244.3 (AFVFPK) 8.62e6 Area 4.22;301179 04Apr2016_CRP_Cone_002 2: MRM of 4 Channels ES+ 565.101 > 347.01 (ESDTSYVSLK) 1.35e7 Area 3.41;477497 04Apr2016_CRP_Cone_002 3: MRM of 2 Channels ES+ 569.135 > 716.9 (GYSIFSYATK) 3.60e6 Area 4.60;119972 04Apr2016_CRP_Cone_002 4: MRM of 3 Channels ES+ 697.287 > 567.1 (QDNEILIFWSK) 4.31e5 Area 5.84 9641 AFVFPK ESDTSYVSLK GYSIFSYATK ACQUITY HSS T3 1.7 µm 2.1 x 50 mm Peptide Precursor Charge State MRM transition Cone Voltage (V) Collision Energy (eV) Product Ion Identification AFVFPK [M+2H]2+ 354.935>244.56 30 15 [2H+]1/y4 and [1H+]1/y2 ESDTSYVSLK [M+2H]2+ 565.101>347.011 15 15 [1H+]1/y3 GYSIFSYATK [M+2H]2+ 569.135>221.58 25 15 [1H+]1/b2 Quantification of Mid-Size Proteins: CRP Tryptic Peptide LC-MS For research use only. Not for use in diagnostic procedures
- 3. ©2015 Waters Corporation 3 CRP: Standard Curve and QC Statistics Excellent Accuracy and Precision CRP digested and extracted in Human Plasma Peptide Curve (µg/mL) Curve (nM) Weighting Linear Fit (R2 ) % Accuracy Range AFVFPK 0.050-100 1.99-3987 1/x2 0.998 93.6-104.4 ESDTSYVSLK 0.050-100 1.99-3987 1/x 0.999 96.8-102.4 Peptide CRP Overspike Concentration (µg/mL) CRP Overspike Concentration (nM) CRP QC Concentration (µg/mL) CRP QC Concentration (nM) Mean (N=3) Calculated Concentration (µg/mL) Mean (N=3) Calculated Concentration (nM) Mean (N=3) % Accuracy %RSD AFVPFK Lot #1 0.000 0.000 0.387 15.420 0.387 15.420 100.0 3.66 0.075 2.990 0.462 18.421 0.481 19.191 104.3 0.60 0.750 29.904 1.137 45.334 1.176 46.889 103.5 2.51 7.500 299.035 7.887 314.466 7.941 316.605 100.7 2.23 75.000 2990.353 75.387 3005.783 74.678 2977.515 99.1 1.25 AFVPFK Lot #2 0.000 0.000 1.162 46.331 1.162 46.331 100.0 4.55 0.075 9.968 1.237 49.321 1.167 46.543 94.4 4.52 0.750 29.904 1.912 76.234 1.750 69.775 91.5 1.15 7.500 299.035 8.662 345.366 8.398 334.853 96.9 2.29 75.000 2990.353 76.162 3036.684 72.141 2876.348 94.7 1.74 AFVPFK Lot #3 0.000 0.000 1.867 74.440 1.867 74.440 100.0 1.20 0.075 9.968 1.942 77.430 1.787 71.250 92.0 3.92 0.750 29.904 2.617 104.343 2.290 91.319 87.5 3.17 7.500 299.035 9.367 373.475 8.527 339.983 91.0 1.29 75.000 2990.353 76.867 3064.793 72.444 2888.429 94.2 13.31 AFVPFK Lot #4 0.000 0.000 18.128 722.778 18.128 722.778 100.0 2.26 0.075 9.968 18.203 725.779 17.612 702.201 96.8 6.47 0.750 29.904 18.878 752.692 17.586 701.178 93.2 1.72 7.500 299.035 25.628 1021.824 23.988 956.421 93.6 5.59 75.000 2990.353 93.128 3713.141 83.284 3320.634 89.4 1.37 For research use only. Not for use in diagnostic procedures
- 4. ©2015 Waters Corporation 4 Quantification of CRP: QC chromatograms Lot #1 75 µg/mL Time 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 13Sep2016_CRPHumanSamples_048 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 2.36e7 Area 4.25 1081770 13Sep2016_CRPHumanSamples_045 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 2.36e7 Area 4.25 117564 13Sep2016_CRPHumanSamples_042 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 2.36e7 Area 4.25 16823 13Sep2016_CRPHumanSamples_039 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 2.36e7 Area 4.25 6767 13Sep2016_CRPHumanSamples_036 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 2.36e7 Area 4.25 5710 Lot #1 7.5 µg/mL Time 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 13Sep2016_CRPHumanSamples_048 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 2.95e6 Area 4.25 1081770 13Sep2016_CRPHumanSamples_045 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 2.95e6 Area 4.25 117564 13Sep2016_CRPHumanSamples_042 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 2.95e6 Area 4.25 16823 13Sep2016_CRPHumanSamples_039 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 2.95e6 Area 4.25 6767 13Sep2016_CRPHumanSamples_036 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 2.95e6 Area 4.25 5710 Blank Lot #1 Time 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 13Sep2016_CRPHumanSamples_048 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 7.38e5 Area 13Sep2016_CRPHumanSamples_045 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 7.38e5 Area 4.25 117564 13Sep2016_CRPHumanSamples_042 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 7.38e5 Area4.25 16823 13Sep2016_CRPHumanSamples_039 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 7.38e5 Area 4.25 6767 13Sep2016_CRPHumanSamples_036 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 7.38e5 Area 4.25 5710 Blank Lot #1 Time 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 2.00 2.25 2.50 2.75 3.00 3.25 3.50 3.75 4.00 4.25 4.50 4.75 5.00 5.25 5.50 5.75 6.00 6.25 6.50 6.75 % 0 100 13Sep2016_CRPHumanSamples_048 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 5.53e5 Area 13Sep2016_CRPHumanSamples_045 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 5.53e5 Area 4.25 117564 13Sep2016_CRPHumanSamples_042 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 5.53e5 Area 4.25 16823 13Sep2016_CRPHumanSamples_039 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 5.53e5 Area 4.25 6767 13Sep2016_CRPHumanSamples_036 Sm (Mn, 2x2) 1: MRM of 2 Channels ES+ 354.708 > 219.113 (AFVFPK) 5.53e5 Area 4.25 5710 AFVFPK Blank Human Plasma 0.075 µg/mL 0.75 µg/mL 75 µg/mL 7.5 µg/mL QC Level 75.387 µg/mL (3005 nM) 7.887 µg/mL (315 nM) 1.137 µg/mL (45 nM) 0.462 µg/mL (18 nM) 0.387 µg/mL (15 nM) Endogenous Concentration 35 µL sample, 2 hour digestion with ProteinWorks eXpress Direct Digest Kit, MCX SPE peptide level clean-up with recovery >90% For research use only. Not for use in diagnostic procedures
- 5. ©2015 Waters Corporation 5 Integrated Microflow LC
- 6. ©2015 Waters Corporation 6 Microflow in Trap and Back-elute Mode Analyte Band Enables higher loading and additional removal of interferences
- 7. ©2015 Waters Corporation 7 Infliximab High Analytical Sensitivity Using Integrated Microflow Time 5.50 5.75 6.00 6.25 6.50 6.75 7.00 7.25 7.50 % 0 100 5.50 5.75 6.00 6.25 6.50 6.75 7.00 7.25 7.50 % 0 100 5.50 5.75 6.00 6.25 6.50 6.75 7.00 7.25 7.50 % 0 100 5.50 5.75 6.00 6.25 6.50 6.75 7.00 7.25 7.50 % 0 100 5.50 5.75 6.00 6.25 6.50 6.75 7.00 7.25 7.50 % 0 100 5.50 5.75 6.00 6.25 6.50 6.75 7.00 7.25 7.50 % 0 100 Std_21Sep_49 MRM of 6 Channels ES+ 469.6 > 603.8 (Remicade SINSATHYAESVK) 4.07e6 Std_21Sep_39 MRM of 6 Channels ES+ 469.6 > 603.8 (Remicade SINSATHYAESVK) 4.70e5 Std_21Sep_29 MRM of 6 Channels ES+ 469.6 > 603.8 (Remicade SINSATHYAESVK) 4.66e4 Std_21Sep_1 MRM of 6 Channels ES+ 469.6 > 603.8 (Remicade SINSATHYAESVK) 1.33e4 Std_21Sep_60 MRM of 6 Channels ES+ 469.6 > 603.8 (Remicade SINSATHYAESVK) 1.14e4 blank_21Sep_8 MRM of 6 Channels ES+ 469.6 > 603.8 (Remicade SINSATHYAESVK) 9.15e3 5.94 7.377.19 blank 10,000 ng/mL 1000 ng/mL 100 ng/mL 10 ng/mL 1 ng/mL For research use only. Not for use in diagnostic procedures
- 8. ©2015 Waters Corporation 8 Total Antibody quantification: TDM-1 35 µL Plasma Time 5.50 5.75 6.00 6.25 6.50 6.75 7.00 7.25 7.50 7.75 % -2 98 5.50 5.75 6.00 6.25 6.50 6.75 7.00 7.25 7.50 7.75 % -2 98 MRM of 17 Channels ES+ 542.77 > 249.16 IYPTNGYTR 1.17e5 Area 4242 MRM of 17 Channels ES+ 542.77 > 249.16 IYPTNGYTR 1.17e5 Area Sample Name Area Height Ret. Time Calc. Conc. %Dev Blank Plasma 10 ng/mL 1834.4 22354 6.78 9.5 -5.3 50 ng/mL 4605.9 82301 6.77 55.8 11.6 100 ng/mL 6834.7 128153 6.77 93.0 -7.0 100 ng/mL 6785.4 122706 6.76 92.2 -7.8 500 ng/mL 25951.6 507460 6.78 412.5 -17.5 500 ng/mL 30172.1 603575 6.78 483.0 -3.4 10000 ng/mL 535725.6 10640577 6.74 8930.6 -10.7 10000 ng/mL 677351.1 13485926 6.76 11297.1 13.0 50000 ng/mL 3185028.5 60066564 6.70 53199.6 6.4 50000 ng/mL 2778577.5 54150816 6.74 46407.9 -7.2 50 ng/mL TDM-1 Blank Plasma
- 9. ©2015 Waters Corporation 9 FTISADTSKNTAYLQMNSLR Miscleavage peptide of T-DM1 blank 50 µg/mL 1 µg/mL 10 µg/mL FTISADTSKNTAYLQMNSLR Miscleavage peptide of T-DM1 For research use only. Not for use in diagnostic procedures
Notas do Editor
- Entire cartridge or what we call ikey is placed into an specially designed ion source. All a user has to do is turn a handle. This handle moved a cam and drives the tile into a series of PEEK fittings. These peek fitting make a face seal very similar to the emitter design. These fittings can handle pressures in excess of 12kpsi. In additonal to the fluidics all of the electronic connections are made as well such as the column heater and electropsray voltage. The tile also has an onboard EPROMM which talks with the LC and logs data such as the number of injections and sample sets.