Beyond Sensitivity Improving the Performance, Productivity ... · Beyond Sensitivity Improving the...

©2011 Waters Corporation 1

Beyond Sensitivity

Improving the Performance, Productivity

and Compliance of the Bioanalytical

Assay Process

EBF, November 17th 2011, Hesperia Towel Hotel

Barcelona

Dr Diego Rodriguez Cabaleiro

Bioanalysis Solution Manager

Waters Marketing Europe


Ultra high sensitivity for

targeted quantification

Information rich qualitative

supporting data

EBF 2010 Xevo TQ-S

ScanWave

Versatility &

Simplicity of operation


Xevo TQ Xevo TQ-S

Larger sampling orifice

How did we increase sensitivity?

Problems associated with a larger sampling orifice:

1) A disperse ion cloud

2) Higher levels of matrix contamination


Ele

ctr

ic F

ield

Diffuse Ion

Cloud

Maximising signal

Maximising robustness

Dealing with a Larger Orifice


Increase in sensitivity!

Fenuron ESI+ 30 7

Metamitron ESI+ 32 15

Acephate ESI+ 27 7

Chlortoluron ESI+ 27 8

Aldicarb ESI+ 27 6

Demeton S Methyl ESI+ 26 9

Phoxim ESI+ 64 19

Kresoxim Methyl ESI+ 64 4

Azinphos Methyl ESI+ 42 6

Azoxystrobin ESI+ 45 4

Dimethoate ESI+ 23 10

Acetamiprid ESI+ 30 28

Fluticasone ESI+ 30 3

Formoterol ESI+ 39 4

Nefadazone ESI+ 28 3

Desmopressin ESI+ 129 25

Salmeterol ESI+ 41 8

Alprazolam ESI+ 21 13

Reserpine ESI+ 25 5

Ibuprofen ESI- 13 16

Prostaglandin E2 ESI- 30 37

Mean Difference 38 11

Relative

Peak Area

Relative

S:NCompound Name

Ionisation

Mode


min

2.20 2.40 2.60 2.80 3.00 3.20 3.40 3.60 3.80 4.00 4.20 4.40

%

0

100

MRM of 5 channels,ES+

504.287 > 293.188

180511_spe_jm_B_V_022 Smooth(SG,2x1)

0.75 0.375

7.162e+0053.68

Fluticasone D3 (1)

3.06

11861.36

623557

2.42 2.772.52

2.82

3.35

min

%

0

100

MRM of 5 channels,ES+

501.28 > 293.185

180511_spe_jm_B_V_022 Smooth(SG,2x1)

0.75 0.375

6.864e+0042.83

2.16

2.07

2.502.432.33

2.25

2.54

2.692.57

Flut 293

3.07

1002.32

44254

2.91

2.95 3.16 3.19

3.283.40

3.49 3.61

4.334.053.94

Compound name: fluticasone

Correlation coefficient: r = 0.999364, r̂ 2 = 0.998729

Calibration curve: 0.663497 * x + 2.14527

Response type: Internal Std ( Ref 2 ), Area * ( IS Conc. / IS Area )

Curve type: Linear, Origin: Exclude, Weighting: 1/x, Axis trans: None

pg/mL0 100 200 300 400 500 600 700 800 900 1000

Response

0

100

200

300

400

500

600

Xevo TQ-S Fluticasone (0.37 pg/mL in Plasma)

Standard

Internal Standard

4 orders of magnitude linear dynamic range


Great success and great feedback for more improvements!

http://www.quotientbioresearch.com/

http://www.yorkbio.com/


Key Customer Challenges in Regulated Bioanalysis

Regulated Bioanalysis

Quality and Consistency

of results

Evolving analytical demands

Sensitive, Robust, meets

assay reqs

Increasing regulatory demands

->Up time

<-Cost per sample


Chemistry

ACQUITY Family

Xevo Family

Informatics

Current Individual Offerings


More Innovation Required

Sensitivity potential of the system not realized

Plenty of room to maximize

Existing platforms in place but with compromise

System Performance

Productivity

Validation & Security

System solution


2011 Introducing Next Generation Regulated Bioanalysis System Solution

• Best in class

Sample Preparation Solutions

• The fastest and most resolving LC without compromise in robustness

ACQUITY UPLC® I-Class

• Different choices for best performance

ACQUITY UPLC Columns

• Most sensitive MS

® TQ-S

• Compliance ready, Interactive workflow-driven data platform

UNIFI

• Installation, maintenance & training

• Compliance services

Service


Sample Preparation Solution

•Improves reproducibility & throughput vs LLE and PPT

•Combined PPT and superior phospholipid removal

•Highest sensitivity, selectivity and reproducibility

•Minimal matrix effects

•HLB and mix-mode & all formats available

•Straight forward method development strategy

•Specific biomolecules application kits

•Reduce sample handling vs classic PPT (no centrifugation…)

•Particulate free filtrate

•Automated, simple method development, easy workflow and high troughput (96 well plates)


Comparison of Phospholipids Remaining after Various Sample Preparation Techniques

MRM of m/z 184-184

Time 0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40 2.60 2.80

%

0

100

0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40 2.60 2.80

%

0

100

0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40 2.60 2.80

%

0

100

0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40 2.60 2.80

%

0

100 184.4 > 184.4 (Lipid 184) 2.00e8

2.88 2.29 2.21

2.10

1.90

2.60 2.78 2.72

184.4 > 184.4 (Lipid 184) 2.00e8

2.80 2.27

1.90

2.62 2.56 2.68

184.4 > 184.4 (Lipid 184) 2.00e8

1.96 1.90 1.77

184.4 > 184.4 (Lipid 184) 2.00e8

2.84 2.21 1.96 1.75 1.42 1.38

1.32

1.63 1.51

PPT

Ostro™

LLE with MTBE

LLE with 5% NH4OH in MTBE


Has minimized dispersion to enhance MS and UV performance

Lowest carryover complementing MS sensitivity and extending MS linear dynamic range

The system’s low dispersion and higher pressure limits, faster cycle-time allows complex separations can be accelerated without compromising chromatographic fidelity

Seamless method trasfer

Robust, proven technology

ACQUITY UPLC I Class


Fastest Possible Scenario (1 mm Column) ACQUITY vs ACQUITY IClass

Instrument Method Name: 30sec_10_95p600mL_90C ACQUITY

0.1

58

0.2

05

0.3

51

0.4

19

0.4

57

0.4

83

0.5

03

AU

0.000

0.022

0.044

0.066

0.088

0.110

Instrument Method Name: 20sec_NO hold 10_95_p904mL_90C

0.0

98

0.1

17

0.1

70

0.2

13

0.2

41 0.2

61

0.2

76

AU

0.000

0.022

0.044

0.066

0.088

0.110

Minutes0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70

Average peak capacity at 5σ = 57

Average peak capacity at 5σ = 70

ACQUITY UPLC

ACQUITY UPLC – I CLASS

Ballistic gradient on 1mm x 50mm BEH C18

Due to the 15k psi limit, with the ACQUITY, the flow rate is 0.6 ml/mn

With the ACQUITY I-Class the flow rate is 0.9 ml/mn


ACQUITY UPLC and Column Choices

Highest resolution and sensitivity (sub 2µm)

Straigh forward method development and transfer

Broad selection of selectivities

Multiple dimensions

Broad range of pH and temperatures

High batch to batch reproducibility

Generic

Polar compounds

Basic compounds

Very polar compounds


Xevo TQ-S

High Performance and information rich data

Stepwave Large increase in sensitivity (all acquisition modes) Designed to be robust to sample matrix contamination

Scanwave Enhanced product ion spectra

RADAR Targeted MRM at the same time as full scan MS

Versatility Ion Sources

Simplicity of Operation

Engineered Simplicity


Rapid MS to MRM Switching


m/z100 200 300 400 500 600 700 800 900 1000

%

0

100

m/z100 200 300 400 500 600 700 800 900 1000

%

0

100253.3

123.0

293.3

397.3

441.4619.6

485.5

329.3

214.1

100.1149.3279.3

251.1

415.3

363.4

546.4

468.6

600.7

Time0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40

%

0

100

0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40

%

9

0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60 1.80 2.00 2.20 2.40

%

2

Targeted compound MRM positive ESI Fluticasone

in plasma

Matrix Profile

MS full scan negative ESI

Matrix Profile

MS full scan positive ESI

Qu

alita

tive

Qu

an

tita

tive

Rapid MS to MRM Switching


Using Dual Scan MRM to Evaluate DBS Card Type To Be Used

DMPK ADMPK A

DMPK BDMPK B

DMPK CDMPK C

Time0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100

0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100

0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100

010210_JoM_DBS_cards_013 2: MS2 ES+ TIC

5.98e91.18

1.030.22

0.32

0.94

1.55

1.421.29

2.411.911.76 2.08 2.24


5.98e9

1.54

0.98

0.22 0.63

1.28

1.40

1.63

2.341.78

1.88 2.052.24

2.41


5.98e91.54

0.23 1.45

1.281.030.90

0.58

2.34

1.73

1.872.07

DMPK ADMPK A

DMPK BDMPK B

DMPK CDMPK C

Time0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100

0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100

0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100


5.98e91.18

1.030.22

0.32

0.94

1.55

1.421.29

2.411.911.76 2.08 2.24


5.98e9

1.54

0.98

0.22 0.63

1.28

1.40

1.63

2.341.78

1.88 2.052.24

2.41


5.98e91.54

0.23 1.45

1.281.030.90

0.58

2.34

1.73

1.872.07

Full scan background

signal

DMPK ADMPK A

DMPK BDMPK B

DMPK CDMPK C

Time0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100

0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100

0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100

020210_JoM_DBS_cards_010 1: MRM of 3 Channels ES+ 309.2 > 281 (Alprazolam)

9.61e5

1.10


9.61e5

1.09


9.61e5

1.09

DMPK ADMPK A

DMPK BDMPK B

DMPK CDMPK C

Time0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100

0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100

0.25 0.50 0.75 1.00 1.25 1.50 1.75 2.00 2.25 2.50 2.75

%

0

100


9.61e5

1.10


9.61e5

1.09


9.61e5

1.09

Analyte Response


UNIFI 1.5: Scientific Information System for Regulated Bioanalysis

Unites our end-to-end system

solution

One software for Chromatography,

MS & Data Management

Workflow based design adjusts to

roles and capabilities of users

Compliance-ready: Designed for

Annex 11 & Part 11 compliance

Scalable, open architecture


Waters Service and Support

Project Management

System Design & Configuration

Installation Services

Training

Instrument Qualification

Software Validation

Repair & Maintenance

Waters Quality parts

Method validation

Method translation


Evolving Analytical Demands - Bioanalysis of Peptides



of results



assay reqs


->Up time

<-Cost per

sample


Therapeutic Peptide Method Development Kit

*= data being generated

Maximum recovery = enhanced sensitivity Minimum matrix effects = selectivity and sensitivity

Peptide pI MW % SPE

Recovery % Matrix

Effects

Octreotide 9.3 1019 88 <10%

Angiotensin II 7.35 1046 82 8%

Desmopressin 8.6 1069 104 <11%

Vasopressin 9.1 1084 100 -3%

Goserelin 7.3 1270 100 -2%

Angiotensin I 7.51 1296 109 *

Somatostatin 10.4 1638 94 *

Neurotensin 8.93 1673 114 6%

Bivalirudin 3.87 2180 100 10%

BNP 12 3464 84 *

Teriparatide 9.1 4118 97 9%

Enfuvirtide 4.06 4492 102 *


MP35N offers a more inert

surface that avoid

adsorptions

Smother internal surface

Updated Tubing Material in I-Class

50x 100x

Conventional SS New MP35N


m/z 400 600 800 1000 1200 1400 1600 1800

%

0

100 Scan ES+

m/z range of Quadrupoles (2048 amu)

Enfuvirtide

MW 4492

Parent ions

— 3+ at m/z 1498

— 4+ at m/z 1124 Requires at least

1500 amu on first quad

1498

1124

m/z 400 600 800 1000 1200 1400 1600

%

0

100

Bivalirudin

— 2 MRM Transitions

o 1091->650

o 1091->1530 • Requires at least 1500 amu on second quad

Singly charged fragments

650

1091 153

0


Increasing Regulatory Demands



of results



assay reqs


->Up time

<-Cost per

sample


Increasing Regulatory Demands: GLP and 21 CFR 11 Compliance

UNIFI

Embedded relational database

Secure server based workgroup architecture

Comprehensive Audit trail

Electronic signature

Built-in Qualification Center

Central repository of work-flow based SOP’s

Fileless connection to Watson LIMS

Structurally Validated calculation of ISR

Automatic calculation and comparison of Matrix Effect

Factor

WATERS SERVICES

Compliance service and consultancy

Instrument Qualification & Software Validation


UNIFI Scientific Information System


Workgroup architecture

<< LABORATORY NETWORK >>

Laboratory Network Appliance

Instrument Control

Server

Presentation Layer

Presentation Layer

Client PC Client PC

Business Layer Database Layer

Instrument Control

Laboratory Network Appliance

Presentation Layer Capture Agent

Client PC

Secure Data Center

Enables Managed Information Sharing & Simplifies Compliance Efforts


N-tier architecture

DATABASE

APPLICATION LAYER ACQUISITION CONTROL

Reporting Data Processing

Event Center Audit Trails

Document Approval Security

Xevo TQ-S

PORTAL CLIENT


Reducing Software Validation Barrier

Present Future Desire

Barrie

rs

SO

FTW

AR

E

VA

LID

ATIO

N

GAMP 5 for COTS Leverage Vendor testing Test Summary document Vendor Quality systems Supplier Assessment

Built in Tools for Qualification Qualification Center to manage IQ and OQ of Software and Instruments Portal installation on Clients Built in Tools for

Verification Development of common Validation Scripts Built in tools to automate running and documenting scripts

Validation Consulting Integrate common scripts into a complete validation package customised to Customer protocols

?

Waters Quality System and Professional Services can significantly reduce the Validation effort


What Will YOU Do With

No compromises. More Choices

Focus on Science

Partnering for the complete solution

Invest for today

and future

Leverage Technology in real time


Questions

Please visit us at the booth

Product Managers

Senior Application Scientists

Software Specialists

Informatics Solution Consultants

Separation Experts

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