chip technology, nanoliters and picoliters - miniaturization of (bio)analytical chemistry methods

... talk given at Lund, 2005

Transcript

1. Chip technology, nanoliters and picoliters – miniaturization of (bio)analytical chemistry methods Andreas Manz I S A S INSTITUTE FOR ANALYTICAL SCIENCES Dortmund and Berlin
2. photo Berlin Dortmund currently 140 staff, 100 in research annual budget EUR 9M
3. organization Prof.Dr.Andreas Manz Prof.Dr.Kay Niemax head miniaturization proteomics metabolomics spectroscopy materials PD Dr.Joachim Franzke Dr.Norbert Jakubow ski Dr.Jörg I. Baumbach PD Dr.Volker K. Deckert Dr.Roland Hergenröder head a.i. head head head head micro plasmas transcription profiling volatile metabolites GC-IMS nano raman synchrotron XRF (PD Dr.Joachim Franzke) Prof. Dr. Philip Day (Dr.Jörg I. Baumbach) (PD Dr.Volker K. Deckert) Alex von Bohlen microfluidic separations ICP-MS molecular imaging functional biotechnology diode laser AS femtosecond laser ablatio (Prof.Dr.Andreas Manz) (Dr.Norbert Jakubow ski) Prof. Dr. Andreas Schmid (Prof.Dr.Kay Niemax) (Prof.Dr.Kay Niemax) x-ray and neutron sources echelle spectrometers* IR ellipsometry* Prof.Dr.Eduardo Greaves Dr.Helmut Becker-Ross PD Dr.Norbert Esser * Berlin-Adlershof
4. vision • identify and quantify all compounds in a mixture („...omics“) • ... as a function of time (monitoring) • ... as a function of space (imaging)
5. vision time space information content 1 times 1 location 1 compound 1d 2d 3d continuously 1/s 1/min „...ome“ complex mixture mixture
6. time space information content proteomics glucose sensor most analytical methods NMR tomography
7. time space vision
11. How can we do it ? What will it cost ? What time does it take ?
12. why miniaturize volume of 1µL 1nL 1pL (1mm)3 (100µm)3 (10µm)3 600,000,000 600,000 600 25 / cm2 2500 / cm2 250 ,000/ cm2 17 min 10s 100ms 1.5 /min / cm2 250 /s / cm2 2,500,000 /s / cm2 # molecules (1nM solution) # volumes In array diffusion time # reactions (diffusion controlled) is a cube of
13. What do we have now? [example 1] • Electrophoresis chips - Caliper, Agilent, Predicant, Hitachi, Shimadzu etc. • mainly used for DNA fragment sizing • protein separations
14. 10 fold miniaturization 100 x faster separation 1000 x smaller volume 10 x lower reagent consumption
15. fluorescence [arb. units] time [s] 0 40 80 120 160 1 2 3 4 5 6 cycle # 7 8 t 7 s synchr. fluorescence [arb. units] time [s] 0 40 80 120 160 1 2 3 4 5 6 cycle # 7 8 t 7 s synchr. fluorescence [arb. units] time [s] 0 40 80 120 160 1 2 3 4 5 6 cycle # 7 8 t 7 s synchr. electrophoresis FITC labeled amino acids D.J.Harrison, K.Flury, K.Seiler, Z.Fan, C.S.Effenhauser, A.Manz, Science 261, 895-897 (1993) C.S.Effenhauser, A.Manz, H.M.Widmer, Anal. Chem. 65, 2637-2642 (1993)
16. electrophoresis double stranded DNA (x 174 HaeIII digest) A.Manz, H.Becker, Transducers 97, Chicago, June 16-19, 1997, Digest of Technical Papers, (ISBN 0-7803-3829-4), 915-918 (1997)
17. nano channels & single molecules 80 x 80 nm channel bulk DNA (-DNA) L.C.Campbell, M.J.Wilkinson, A.Manz, P.Camilleri, C.J.Humphreys, Lab Chip 4, 225-229 (2004)
18. Agilent 2100 Bioanalyzer electrophoresis
19. Mass spec spraying needle, needle assembly & fittings Nano LC Column Enrichment column, capillaries, fittings, frits HV ESI contact RF tag chip based LC/MS by Agilent courtesy of Tom A.van de Goor, Agilent, Santa Clara CA
20. What do we have now? [example 2] • Reactor chips - Upchurch, etc. • Mainly used for solvent gradients in chromatography • Chemical synthesis • Bioassays
21. chemical reactor F.G.Bessoth, A.J.de Mello, A.Manz, Anal. Comm. 36, 213-215 (1999)
23. Continuous flow method
24. 0 ms 6 ms chemical reactor
25. Upchurch Scientific chemical reactor
26. Inlet capillaries Syringes Rheodyne injection valve Injection loop outlet capillary Micromixer chip / PTFE interface chemical reactor
27. N+ H H R1 R2 Cl- H H O MeOH N+ Cl- H2 O N R3 R4 R2 R1 C R1 N R2 N R4 R3 H2 O R1 N R2 N R4 R3 O R1/R2 = -CH2 (CH2 )3 CH2 - Piperidine hydrochloride + + Piperidinium cation + R3/R4 = -CH2 (CH2 )4 CH2 - Cyclohexyl isocyanide Nitrilium intermediate -Dialkylacetamide Formaldehyde N-Cyclohexyl-2-piperidin-1-yl-acetamide (1) (2) (3) (4) (5) (6) Multicomponent Chemistries: The Ugi Reaction 0oC
28. Simultaneous Observation of Reactants, Intermediates, Products and By-products 20 mLmin-1 50 nL injection loop Room temperature M.C.Mitchell, V.Spikmans, A.Manz, A.J.de Mello J.Chem.Soc., Perkin Trans.1, 2001, 514-518 (2001) educt intermediate side product main product intermediate
29. horseradish peroxidase assay flow rate dependence of signal <11.5s <570ms 50 ng/mL HRP
30. horseradish peroxidase assay calibration curves
31. Ubiquitin Native/A state Native state A state Methanol pD=2
32. Set-up for NMR measurement Syringe pumps NMR Micromixer Detection coil (200mm i.d.) 250mm i.d. 75mm i.d.
33. Picture of detection coil Sweedler group, Univ. of Illinois Reservoir Capillary 1cm
34. NMR set-up Syringe pump 3m Capillary NMR NMR Sweedler group, Univ. of Illinois
35. 10 mL/min (24sec) 40 mL/min (6sec) His68 Tyr59 N N N N A A A A M.Kakuta, D.A.Jayawickrama, A.M.Wolters, A.Manz, J.V.Sweedler, Anal.Chem. 75, 956-960 (2003)
36. 1643 cm-1 1661 cm-1 1671cm-1 wavenumber [cm-1] 1600 1620 1640 1660 1680 1700 arbitrary units -2e-5 0 2e-5 4e-5 6e-5 2nd derivative A Result (FT-IR) wavenumber [cm-1] 1600 1620 1640 1660 1680 1700 arbitrary units -4e-5 -2e-5 0 2e-5 4e-5 6e-5 8e-5 1e-4 absorbance [AU] 0.000 0.005 0.010 0.015 0.020 0.025 ubiquitin (native) ubiquitin (mixed) absorbance 2nd derivative M.Kakuta, P.Hinsmann, A.Manz, B.Lendl, Lab Chip 3, 82-85 (2003) N N
37. Post-column reactor: µLC-Chip-MS Set-up 25cm x 100µm µLC column Injector Waste LC pumping system 35cm x 50µm TOF MS Electrospray Micromixer chip Syringe pump TMPP+ / TEA CMPI UV detector
38. µLC-Chip-MS Set-up Electrospray needle High voltage Nebulising gas
39. Br - (MeO)3 Ph P + Ph(OMe)3 (MeO)3 Ph N H NH2 O R H O + Br - (MeO)3 Ph P + Ph(OMe)3 (MeO)3 Ph N H N O H R Acetic acid, 30 mins, sonicate Aldehyde 4-Hydrazino-4-oxobutyl-TMPP+Br- Hydrazino derivative of aldehyde Reaction of Aldehydes (and ketones) with 4-hydrazino-4-oxobutyl-TMPP
40. Gradient µLC-chip-MS of Ketones/Aldehydes Column 10cm long 200µm i.d. 5µm Hypersil C18 Ultra High Purity Elite Separation 5 minute gradient from 0 to 90% acetonitrile in water, containing 0.1% formic acid Injection 50nL Reagents all 1.0 mM at 1µL/min MS Micromass Q-Tof II Ketones/Aldehydes Products Cyclohexanone Valeraldehyde Cyclohexane carboxaldehyde Heptaldehyde V.Spikmans, S.J.Lane, B.Leavens, A.Manz, N.W.Smith, Rapid Commun. Mass Spectrom. 16, 1377-1388 (2002)
41. Gradient µLC-chip-MS of Amines – isotope labeling analysis Methylheptylamine Methyloctylamine Methyldecylamine Methylhexylamine Products Accurate Mass Difference Isotopes: 2.012 Only characteristic pattern is necessary, not molecular weight
42. Sequential DNA hybridization • Inject small volume plugs of probe DNA oligomers • Mix within ms with target DNA • Observe hybridization reaction as the plug moves downstreams
43. DNA hybridisation assay Intercalating dye alone low DNA oligomers low Oligomer dimers medium dsDNA high
44. Influence of DNA Sequence on Fluorescence Levels Order: matching, 1 mismatch, 2 mismatches, 5 mismatches Order: matching, 1 mismatch, 2 mismatches Sequence-dependent responses from two different experiments.
45. Quick Decision: Exploiting Photobleaching Effects M.Heule, A.Manz, Lab Chip 4, 506-511 (2004)
46. microfluidic DNA assays • 1 second to decision • no complicated surface chemistry • sensitivity 100-200nM • could be competing with DNA arrays
47. What would I address? [example ] protein separations by free-flow electrophoresis … isoelectric focusing
48. free-flow electrophoresis - proteins + -
49. IEF chip • 36 x 20 um inlet channels • 72 x 20 um outlet channels • each side 108 x 4 um channels • separation bed 12.2 x 4.1 mm – 15,552 posts – 30 x 30 um
50. free-flow electrophoresis
51. very fast electrophoresis C.-X.Zhang, A.Manz, Anal. Chem. 75, 5759-5766 (2003)
52. fluorescence [arb. units] time [s] 0 40 80 120 160 1 2 3 4 5 6 cycle # 7 8 t 7 s synchr. fluorescence [arb. units] time [s] 0 40 80 120 160 1 2 3 4 5 6 cycle # 7 8 t 7 s synchr. fluorescence [arb. units] time [s] 0 40 80 120 160 1 2 3 4 5 6 cycle # 7 8 t 7 s synchr. comparison FFE CE
53. IEF FFE Isoelectric focusing using free-flow electrophoresis
54. IEF proof of principle 12 mm 0 mm 4 mm 4 mm = 500 ms angiotensin I, 1.75 kV, 10 uL/min Y.Xu, C.X.Zhang, A.Manz, Lab Chip 3, 224-227 (2003)
55. IEF - proteins angiotensin I, 1.75 kV, 10 uL/min 400ms
56. IEF - IGF-1 10μm 4mm
57. IEF chip • volume 240 nL plus wells • at 10 uL/min – 1.4 seconds (time to information) • preconcentration 100 - 400x
59. Chip technology, miniaturization of (bio)analytical chemistry methods Andreas Manz I S A S INSTITUTE FOR ANALYTICAL SCIENCES Dortmund and Berlin
60. Andreas Manz I S A S INSTITUTE FOR ANALYTICAL SCIENCES Dortmund and Berlin … fun stuff from the Manz’ new lab
61. T.Vilkner, A.Shivji, A.Manz, Lab Chip 5, 140-145 (2005)
62. dry powder dispenser T.Vilkner, A.Shivji, A.Manz, Lab Chip 5, 140-145 (2005) principle: fluidized bed
63. dry powder dispenser T.Vilkner, A.Shivji, A.Manz, Lab Chip 5, 140-145 (2005) principle: fluidized bed
64. dry powder dispenser T.Vilkner, A.Shivji, A.Manz, Lab Chip 5, 140-145 (2005) reproducible injections of 1 – 50 mg of dry powder (non-cohesive)
65. MALD for bacterial spore disruption O.Hofmann, K.Murray, A.S.Wilkinson, T.Cox, A.Manz, Lab Chip 5, in press 2005 Matrix Assisted Laser Desorption
66. MALD for bacterial spore disruption O.Hofmann, K.Murray, A.S.Wilkinson, T.Cox, A.Manz, Lab Chip 5, in press 2005 6mW nitrogen laser 337nm 4ns pulse width 75kW peak power
67. MALD for bacterial spore disruption O.Hofmann, K.Murray, A.S.Wilkinson, T.Cox, A.Manz, Lab Chip 5, in press 2005 before Bacillus globigii spores 3-hydroxypicolinic acid after
68. Np XRF chip E.Greaves, A.Manz, Lab Chip 5, in press 2005 Am 241 95 Np 237 93 458 years α emitter 900 nCi
69. Np XRF chip E.Greaves, A.Manz, Lab Chip 5, in press 2005 h
70. Np XRF chip E.Greaves, A.Manz, Lab Chip 5, in press 2005 Sn Zn Ni
71. Vac. gauge Pump HV supply ADC HV Amp Pre HP Ge PC CPU S100 HV Leak NIM bin A.- B.- X-ray chip E.Greaves, A.Manz, Lab Chip 5, in press 2005
72. X-ray chip E.Greaves, A.Manz, Lab Chip 5, in press 2005 0.1-10 mtorr 200-600 nA 4 mW
73. cyclic separations
76. chromatography like
77. injection detection speed measurement
79. injection detection speed measurement
80. filling the sample loop… here is the chromatographer ! and, injectioooon !!! … some irreversible processes …
85. linear 1d separations injection detection
86. linear 1d separations injection of small sample volume single point detection resolution depends on pressure applied, or voltage applied
87. linear 1d separations today’s liquid chromatography is at upper pressure limit today’s electrophoresis is at voltage limit
88. how to overcome? somehow keeping compound of interest in same position
89. 1937 A.Tiselius flow-counterbalanced electrophoresis Trans. Fraday Soc. 1937, 33, 524-531 1990 S.C.Lee et al EOF-counterbalanced CE 1994 C.T.Culbertson et al flow-counterbalanced CE
90. how to overcome? mass spectrometry: time-of-flight MS vs. cyclotron MS particle accelerators: linear accelerator vs. synchrotron
91. cyclic separation is not new… 1962 direct pumping recycling LC: Porath, J., Bennich, H., Arch. Biochem. Biophys.1962, Suppl.1 , 152–156.
92. 1962 direct pumping recycling LC: column inj det p valve
93. cyclic separation is not new… 1971 alternate pumping recycling LC: Biesenberger, J. A., Tan, M., Duvdevani, I., Maurer, T., J. Polym.Sci.B Polym.Lett.1971, 9 , 353–357.
94. 1971 alternate pumping recycling LC: column inj det p valve column
95. cyclic separation is not new… 1993 synchronized cyclic CE: Burggraf, N., Manz, A., Effenhauser, C. S., Verpoorte, E., De Rooij, N. F., Widmer, H. M., HRC-J.High Resolut.Chromatogr.1993, 16 , 594–596.
96. 1993 synchronized cyclic CE: + -
97. 1993 synchronized cyclic CE: + -
98. 1993 synchronized cyclic CE: + -
99. cyclic separation is not new… 2001 electrophoretron: Choi, J. G., Kim, M., Dadoo, R., Zare, R. N., J.Chromatogr.A, 2001, 924 , 53–58.
100. 2001 electrophoretron: + -
101. advantages plate number N and resolution Rs are increasing over time same driving force is used multiple times
102. problems peak capacity is going towards 0 many sample components are eliminated pump or valve volume decreases N time
103. vA vB detection window separation channel vsolution • in principle infinitely long separation column • small driving force: determined by circle circumference • resolution proportional to the square root of time Fourier transform chromatography
104. issues constant cross-section around cycle (pump?) high flow velocity for small channel diameters (pump?) overtaking of sample components (detection?) panic
105. overtaking of sample components (detection?)
106. Shah Convolution - FT- Detection H. J. Crabtree, M. U. Kopp and A. Manz, Anal. Chem., 1999, 71,
108. wavelet transform J.C.T.Eijkel, Y.C.Kwok, A.Manz, Lab Chip, 2001, 1, 122.
109. 7 Hz 14 Hz wavelet transform
110. vA vB detection window separation channel vsolution Fourier transform chromatography
111. constant cross-section around cycle (pump?)
112. pumping for cyclic LC separation electrohydrodynamic magnetohydrodynamic AC electroosmotic shear panic
113. magnetohydrodynamic Eijkel, J. C. T., Dalton, C., Hayden, C. J., Burt, J. P. H., Manz, A., Sens.Actuators B 2003, 92 , 215–221.
114. Debesset, S., Hayden, C. J., Dalton, C., Eijkel, J. C. T., Manz, A., Lab Chip 2004, 4 ,396-400. AC electroosmotic
115. shear flow pumping G. Desmet and G. V. Baron, J. Chromatogr. A, 1999, 855(1), 57.
116. shear flow pumping movement of plate v fluid velocity v/2 movement of plate = 0 G. Desmet and G. V. Baron, J. Chromatogr. A, 1999, 855(1), 57.
117. shear flow pumping movement of plate v fluid velocity v/2 movement of plate = 0
118. shear flow pumping movement of plate v fluid velocity v/2 movement of plate = 0
119. shear flow pumping non-retained v/2 retained = 0 stationary phase
120. shear flow pumping non-retained v/2 retained = 0
121. shear flow pumping non-retained v/2 retained = 0
122. shear flow pumping retained v non-retained v/2 stationary phase
123. shear flow pumping retained v non-retained v/2
124. shear flow pumping retained v non-retained v/2
125. shear flow pumping very high speeds possible nm gaps should lead to high N main problem: the end of the plate is the end of the pumping (no more than N=10,000 shown so far)
126. chip design channel width 2mm, depth 15um channel circumference 62.8mm
130. Fourier transform chromatography shear flow pumping Rotation stage Loading direction Chips Stationary clamp Injection pump
131. stationary phase Supershere 60 RP-8 particles tri-methylopropane-trimethacrylate
132. fully retained sample, raw data u = 1.5mm/s (Péclet no = 14) mobile phase methanol/water 1:1 sample coumarin dyes
133. fully retained sample, FT time scale
134. fully retained sample, FT chromatographic window panic
135. non-retained fully retained panic
136. non-retained fully retained panic
137. non-retained fully retained panic
138. non-retained fully retained, or overtone retained …a separation…?
140. conclusions • cyclic separation can be done without sample loss • plate numbers and resolution increase with time • shear flow pumping most promising • deconvolution of detection signal by Fourier or wavelet transform
141. open questions • which signal deconvolution is best? • what dimensions are optimal for performance (theory)? • is there a simultaneous use for two independent stationary phases? • how about gradient elution?
142. Acknowledgment Oliver Hofmann Torsten Vilkner Xin Yang Eduardo Greaves, Prof. Jan Eijkel Sebastien Debesset Dirk Janasek Gareth Jenkins Joachim Franzke Qinetiq, UK Pfizer, UK Smiths Detection, UK Hitachi Ltd, Japan Universidad Simon Bolivar, Caracas, Venezuela Mercator professorship, Germany
143. any questions?