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chip-based microfluidic methods for analytical chemistry

andreas manz
September 25, 2008

chip-based microfluidic methods for analytical chemistry

... talk given at Bologna, September 2008.

andreas manz

September 25, 2008
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  1. Chip based microfluidic methods
    Chip-based microfluidic methods
    for analytical chemistry
    y y
    Andreas Manz
    U K
    U.K.

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  2. microfabrication, 1979

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  3. View Slide

  4. glass glass device, 1747

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  5. 747
    y 17
    logy
    hnol
    tech
    not
    na

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  6. micro-
    micro-
    fluidics,
    ,
    1747

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  7. many things start off with sophisticated
    many things start off with sophisticated
    manual methods
    d l i t f l f l
    develop into fool-proof general
    methods
    methods
    finally, crude power dominates the field

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  8. algorithm to calculate square root of a
    algorithm to calculate square root of a
    number

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  9. l ith i t bl lid l
    logarithmic tables, slide rules

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  10. algorithm to calculate square root of a
    algorithm to calculate square root of a
    number
    l ith i t bl lid l
    logarithmic tables, slide rules
    PC

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  11. information
    content
    vision
    content
    „...ome“
    complex mixture
    mixture
    1 d ti l
    mixture
    time
    1 times
    1 location
    1 compound continuously
    1/s
    1/min
    1d
    2d
    3d
    space
    3d

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  12. information
    content
    content
    proteomics
    glucose sensor
    most analytical
    methods
    time
    methods
    NMR tomography
    space

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  13. information
    content
    content
    vision
    time
    space

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  14. “l b hi ”
    “lab on a chip”
    microfabrication
    microfabrication
    microfluidics
    μTAS
    miniaturized total analysis systems

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  15. s
    ip”
    cs
    pers
    a chi
    uidic
    0 pap
    on a
    roflu
    0,000
    “lab
    micr
    a. 10

    ca

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  16. microfluidics / scaling laws
    t i k
    trick:
    every existing chemistry will work the
    every existing chemistry will work the
    same on small as on large scale
    g

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  17. scaling laws for microwell plate
    scaling laws for microwell plate
    volume of 1µL 1nL 1pL
    (1mm)3 (100µm)3 (10µm)3
    is a cube of
    600,000,000 600,000 600
    # molecules
    (1nM solution)
    25 / cm2 2500 / cm2 250 ,000/ cm2
    # volumes
    In array
    17 min 10s 100ms
    diffusion time
    1.5 /min / cm2 250 /s / cm2 2,500,000 /s / cm2
    # reactions
    (diffusion controlled)
    (diffusion controlled)

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  19. detection in small volumes is an issue
    detection in small volumes is an issue
    going nano is getting worse
    g g g g

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  20. microfluidics / scaling laws
    microfluidics / scaling laws
    trick works for:
    h i l ti
    chemical reaction
    separation
    separation
    dilution series
    etc

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  21. for more information on the topic
    for more information on the topic
    micro TAS conference
    S Di USA 2008
    San Diego, USA 2008
    Cheju, Korea 2009
    impact factor 5.8
    1,000 attendees annually
    p f
    Reviews on Micro total analysis systems
    in Anal.Chem. 2002, 2004, 2006 and 2008
    cited over 1,900 times

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  22. 1
    part 1
    p

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  23. electrophoresis
    p

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  24. scaling laws
    g

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  25. 10 fold miniaturization
    100 x faster separation
    p
    1000 x smaller volume
    10 x lower reagent consumption

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  26. electrophoresis
    FITC l b l d i id
    FITC labeled amino acids
    c
    e [
    a
    r b
    .
    u n
    i
    t
    s ]
    1 2
    3
    4 c
    y
    c l
    e
    #
    t
    7 s
    s
    y
    n
    c
    h
    r
    .
    c
    e [
    a
    r b
    .
    u n
    i
    t
    s ]
    1 2
    3
    4 c
    y
    c l
    e
    #
    t
    7 s
    s
    y
    n
    c
    h
    r
    .
    c
    e [
    a
    r b
    .
    u n
    i
    t
    s ]
    1 2
    3
    4 c
    y
    c l
    e
    #
    t
    7 s
    s
    y
    n
    c
    h
    r
    .
    f l
    u
    o r
    e
    s
    c e
    n
    0 4
    0 8 0 1
    2
    0 1
    6 0
    5
    6
    7 8
    f l
    u
    o r
    e
    s
    c e
    n
    0 4
    0 8 0 1
    2
    0 1
    6 0
    5
    6
    7 8
    f l
    u
    o r
    e
    s
    c e
    n
    0 4
    0 8 0 1
    2
    0 1
    6 0
    5
    6
    7 8
    D J H i K Fl K S il Z F C S Eff h A M S i 261 895 897
    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)

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  27. publications per month
    publications per month
    citing 2100 bioanalyzer
    200
    220
    C
    140
    160
    180
    Courtesy
    80
    100
    120
    y of Agi
    40
    60
    80
    ilent W
    0
    20
    -00
    -00
    -01
    -01
    -02
    -02
    -03
    -03
    -04
    -04
    -05
    -05
    -06
    -06
    Waldbro
    Jan
    Jul
    Jan
    Jul
    Jan
    Jul
    Jan
    Jul
    Jan
    Jul
    Jan
    Jul
    Jan
    Jul
    nn

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  28. 2
    part 2
    p

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  29. serial to parallel converter
    SERIAL
    1 2 3 4
    CONVERTER ?
    PARALEL
    1
    2
    PARALEL
    2
    3
    4

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  30. SERIAL
    1
    1
    CONVERTER
    1
    PARALEL
    2
    3
    3
    4

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  31. 17 SEPARATION CHANNELS
    INJECTION CHANNEL
    INJECTION CHANNEL

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  32. View Slide

  33. View Slide

  34. View Slide

  35. double stranded DNA separation

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  36. reaction with intercalating dye

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  37. View Slide

  38. x
    x
    x
    x
    x x
    x
    x x
    x
    x
    x
    x
    x x
    x
    x
    double stranded DNA SYBR green
    complex
    [fluorescing]
    ration
    DNA is slowing down at
    moving front of SYBR green
    SYBR green is slowing down at
    moving front of DNA
    concentr
    length of plug
    orescence
    fluo
    length of plug

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  39. 3
    part 3
    p

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  40. no scaling laws
    for cell biology
    hi
    on chip

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  41. size similarity of cells and
    microchannels
    ll h t “f l h ”
    cells have to “feel happy”

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  42. timescale 1-2 weeks

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  43. Start Position & Current Situation
    in Stem Cell Research
    H i
    Brain
    Blood
    vessels
    Ear
    Tooth
    1. Take stem cells from any origin
    Hair
    Lung
    Heart
    Pancraes
    Liver
    Kidney
    Kidney
    Cartilage
    2. Induce differentiation by:
    Soluble
    Immobi-
    lised
    C ge
    Bone
    factors
    lised
    factors
    Muscle
    3. Count and find model cell
    type to test the device
    43
    Cell Programming by
    Nanoscaled Devices

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  44. Problem 1: The Difficulty in Comparing Results
    St ll (SC)
    Inner cell group
    In vitro ESC
    Stem cell (SC) sources
    tial of
    ntiation
    higher embryonic
    adult
    Single ESC
    In vitro ESC
    (no. of passage?)
    In vitro adult
    SC (passage?)
    Potent
    differen
    lower
    adult
    on
    low
    Morphology only
    Specific marker (images only)
    Statistics
    ifferentiati
    Specific markers (quantitatively)
    Proteome-
    charac-
    Quality of d
    Functionality Functionality
    and cell
    interaction
    Complete
    epigenetic
    h t i ti
    terisation
    Q
    high
    characterisation
    Long-term stable
    implantation
    44
    Cell Programming by
    Nanoscaled Devices

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  45. Problem 2: Statistical Significance of Differentiation
    90
    100%
    1. Spontaneous differentiation
    depends on many factors. Not
    constant!
    2 No 100% no 0% differentiation
    ]
    70
    80
    90
    Embryonic SC Adult SC
    2. No 100%, no 0% differentiation.
    3. Suppressing any differentiation in
    the case of ESC only.
    ells in [%
    50
    60
    70
    differentiation
    fferentiation
    4. Frequently many factors are
    changed simultaneously.
    ntiated ce
    n
    tion
    30
    40
    pontaneous d
    Induced di
    5. Isotrope versus cluster differentiation.
    of differe
    differentiation
    ed differentiat
    10
    20
    Sp
    erentiation
    Amount o
    Spontaneous
    Induce
    ?
    0
    ppressed diffe
    S
    ?
    45
    Cell Programming by
    Nanoscaled Devices
    Sup

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  46. Problem 3:
    Differentiation is not in vivo Differentiation
    I it lt f t ll
    In vitro-culture of stem cells
    A li i f
    Application of
    factors
    SPARC
    Variant 1:
    X
    Variant 2: Variant n:
    SPARC + X
    Variant 3:
    X + Y
    SPARC X SPARC + X
    + Y
    X + Y



    Actual single factor induction of cell differentiation!
    Application of different media!
    pp
    Unphysiological high concentration of factors!
    In addition undefined substances (e.g. FCS, Trypsin …)!
    46
    Cell Programming by
    Nanoscaled Devices

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  47. SC lines installed and investigated in CellPROM
    in vivo
    Embryogenese:
    highest accuracy in cell
    highest accuracy in cell
    location and differentiation
    in space and time!
    47
    Cell Programming by
    Nanoscaled Devices

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  48. Hundreds of Nanoscapes & Extra Equipment
    48
    Cell Programming by
    Nanoscaled Devices

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  49. MagnaLab: Long-term Cell Cultivation & Differentiation
    Device
    49
    Cell Programming by
    Nanoscaled Devices

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  50. MagnaLab - Cell Cultivation on Carriers over Weeks
    Highly parallel
    NANOSCAPES
    Variable s rface mediated and
    NANOSCAPES
    Variable surface-mediated and
    soluble factor application
    50
    Cell Programming by
    Nanoscaled Devices

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  51. Results of CellPROM
    P i i l f M L b
    Principle of MagnaLab
    Bild MagnaLab
    20
    Carrier
    120
    Microcarriers
    in one
    cultivation
    unit
    20
    Carrier
    20
    Carrier
    unit
    20 Carrier
    20 Carrier
    20
    Carrier
    Cultivation for more than 20 days!
    ! Inlet and outlet tubes removed !
    51
    Cell Programming by
    Nanoscaled Devices
    Cultivation for more than 20 days!

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  52. R lt f C llPROM
    Results of CellPROM
    Cultivation for more than 20 days!
    52
    Cell Programming by
    Nanoscaled Devices
    Cultivation for more than 20 days!

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  53. Principle of Microcarrier-Multichannel
    Cultivation
    Nanoscape immobilised factor
    Spontaneous differentiation
    Soluble factor
    Spontaneous differentiation
    BSA BSA
    Suppressed differentiation
    Suppressed differentiation
    LIF
    Induced differentiation
    LIF
    Induced differentiation Induced differentiation
    SPARC SPARC
    53
    Cell Programming by
    Nanoscaled Devices

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  54. Beating Cardiomyocyte Clusters Stem Cell
    Colonies
    X = Number of beating (synchronised) clusters/cm2 Y = Number of stem cell clusters/cm2
    50 up to 100 synchronized
    54
    Cell Programming by
    Nanoscaled Devices
    50 up to 100 synchronized
    cardiomyocytes

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  55. work in progress
    …work in progress
    mm scale device handling
    m scale channels
    m scale channels
    nm scale surface chemistry
    nm scale surface chemistry
    5-15 days time scale

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  56. part 4
    part 4
    panic

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  57. • soft matter for microfluidics ?
    biocompatible
    biocompatible
    self assembly
    potentially low cost

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  58. View Slide

  59. vesicle production
    PDMS
    p
    PDMS
    Si
    PDMS
    Si
    PDMS

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  60. vesicle production
    p
    PDMS
    Si
    PDMS 100 µm
    2 µm

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  61. vesicle production
    p
    flow direction
    100 µm
    100 µm
    side view side view
    fluorescence image

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  62. Formation of vesicles
    100 µm 100 µm 100 µm
    1 0 0
    1 2 0
    1 4 0
    4 0
    6 0
    8 0
    #
    100 µm 2 4 6 8 1 0 1 2
    0
    2 0
    d ia m e te r (µ m )
    (µ )
    Lipid: DLPC (16:0 Phosphocholine)
    dye: DiI-C18

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  63. formation of vesicles
    P.S.Dittrich, M.Heule, P.Renaud, A.Manz
    Lab Chip 6 488-493 (2006) formation of vesicles
    Lab Chip 6, 488 493 (2006)
    i fl
    increase flow
    increase backside
    pressure
    pressure

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  64. formation of vesicle tubes

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  65. formation of vesicle tubes

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  66. formation of vesicle tubes

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  67. Stopping the flow
    P.S.Dittrich, M.Heule, P.Renaud, A.Manz
    Lab Chip 6, 488-493 (2006)

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  68. Formation of helices
    50 µm
    50 µm
    P.S.Dittrich, M.Heule, P.Renaud, A.Manz
    Lab Chip 6, 488-493 (2006)

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  69. i
    panic

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  70. View Slide

  71. View Slide

  72. View Slide

  73. Part 5 fun
    panic

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  74. hottest issues:
    1 ll bi l t
    1.cell biology support
    2 widening gap between academic
    2.widening gap between academic
    research and industry needs
    y
    3.expiry of microfluidic patents in
    coming few years

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  75. acknowledgements
    g
    Dr Petra Dittrich
    Dr.Petra Dittrich
    Dr.Jonathan West
    Li Ch
    Prof.Günter Fuhr, St. Ingbert
    Lin Chen
    Helke Reinhardt
    g
    Dr.Daniel Schmidt, St.Ingbert
    Prof Claude Leclerq Paris
    Kaoru Tachikawa
    Claus Schumann
    Prof.Claude Leclerq, Paris
    Dr.Richard Loman, Paris
    Prof Philippe Renaud Lausanne
    Dr.Joachim Franzke
    Prof Philip Day
    Prof.Philippe Renaud, Lausanne
    Dr.Martin Heule, Lausanne
    i i
    Prof.Philip Day
    Ying Cai
    P f K i hi Oh
    Dr.Luc Bousse, Mountain View
    Prof.Ken-ichi Ohno

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  76. h d
    the end

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