Jeremy Colson, Boston University 1

Resonant waveguide grating biosensor for living cell sensing

Ye Fang, Ann M. Ferrie, Norman H. Fontaine, John Mauro, and Jitendra Balakrishnan

Biochemical Technologies, Science and Technology Division, Corning Incorporated

Biophysical Journal, June 2006

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Presentation Outline

• Motivation• Background: assays, RWGs• Methods

– Vertical and horizontal mass distributions• Results

– Cell adhesion and spreading– Cell detachment

• Conclusion• Where are they now?

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Jeremy Colson, Boston University 4

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Assays

• Procedure to determine the concentration of a component part of a mixture

• Cell-based– More complex, less specific– Useful for functional information

• Pathway activation• Toxicity• Phenotypic responses

– Need for label-free

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Resonant Waveguide Gratings & MRCAT

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Mass redistribution detection method

• Intensity as a function on incident angle– Resonant peaks

• Vertical mass redistribution– Shift in resonant peak

• Lateral mass redistribution– Changes in peak shape (PWHM)

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Effective refractive index

1 response unit = 5.82x10-4

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Vertical mass redistribution

ii

ii

CnCnn

0

cc

nnNN

Specific refractive index incrementα = .0018 per 100ml/g (protein) .0016 per 100ml/g (other, Na)

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Comments on ΔN

1. Primarily sensitive to vertical mass redistribution (DMR)2. Directly a function of changes in protein concentration due

to protein relocation (rather than ion mobilization) mediated by a stimulation

3. Relocation of a target or complex of certain mass near the sensor surface contributes more to the overall response than those further away

4. Optical signature is an integrated signal that is a sum of contributions from mass redistribution occurring at different distances away from sensor surface

cc

nnNN

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Horizontal mass movement

• Lateral inhomogeneity does not affect refractive index

• Lateral inhomogeneity does affect shape of resonant peaks

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Cell adhesion and spreading• Human epidermoid

carcinoma cells (A431) in 5% FBS

1. At room temp adhesion not optimal

2. Cells interact with surface through multiple steps

3. Spreading step increases the mass w/in sensing volume

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Cell spreading inhibitor added

• 100nM vincristine• Reduced kinetics of

cell spreading• Initial steps primarily

affected• Are the effects of

vincristine limited to first 14 hours?

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Cell detachment

• Trypsin – pancreatic serine protease with substrate specificity based on positively charged lysine and argenine side chains

• Used for cell detachment

• A431 cells, 95% confluency

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Low-doses: cell signaling• Presence of P-DMR:

cell signaling• Slight N-DMR:

insignificant cell detachment

• => activation of endogenous protease-activated receptors that lead to typical Gq signaling

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Conclusions

• Optical signatures are integrated responses that can be used to examine cells in native environments label-free

• Systematic investigation of cell processes– Adhesion– Detachment– Cell signaling: EGFR and Bradykinin B2

receptors

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Back to Epic…

Jeremy Colson, Boston University 18

Thank you!