Measuring Zeta Potential of M-Cells in Mucosal Epithelia

Abstract

Specialized microfold cells (M-cells) play an important role in transport of antigens across the mucosal lymphoid tissues to initiate an immune response. Understanding the mechanism by which M-cells bind antigens can provide insight to mucosal vaccination methods. Previous research has shown that surface charge of bacteria and potential delivery vehicles appears to play a significant role in M-cell selection. Thus, measuring electrostatic and zeta potential at the surface of shear between tissue and bacterial interaction remains as a main goal of our research studies. In this research, the streaming potential of M-cells cultured on membrane surfaces was determined. A streaming potential device was modified and tested to accommodate the membranes. The device was tested known samples consisting of Bovine Serum Albumin (BSA) adsorbed on composite regenerated cellulose (CRC) or polyethersulfone (PES) membranes with a molecular weight cutoff of 30 kDa. Then zeta potential was calculated by Helmholtz-Smoluchowski equation.

Nilufer Nurinovaa, David Lob and Victor G. J. Rodgersb

a Mathematics & CS Department, Tennessee Technological Universityb Bioengineering and Biomedical Sciences, University of California, Riverside

Mucosal M-Cell TargetingPeyer’s Patch M-Cells

Images courtesy of David Lo

Membranes Used for Electrostatic Potential Test

Composite Regenerated Cellulose Membrane (CRC)

Surface properties• Not charged • Unaffected by pH

Polyethersulfone (PES)

Surface properties• Negatively charged at

most pH conditions

Schematic diagram of the device for streaming potential test

[1]

• Protein coated CRC and PES membranes were created by soaking in BSA solutions overnight

• Non-protein coated CRC and PES membranes were created by soaking in nanopure water overnight

• Each membrane was put into the device to test for the streaming potential using 0.15M NaCl with varying pH

• The changing streaming potential (ΔΨ) was recorded for varying hydraulic pressures (ΔP)

• Zeta potential (ζ) was calculated by Helmholtz-Smoluchowski equation[2]

0

0

r

P

ε0 - Permittivity of free space εr - Relative dielectric constant of the solvent µ - Viscosity of the solutionΛ0 - Conductivity of the electrolyte solution

Acknowledgements

Zeta Potential Results of Protein Coated CRC and PES Membranes

Conclusions and Future Work

Membrane Coating Zeta Potential (mV) - pH 7.0

CRC BSA+NaCl 19 + 5.86

CRC BSA+H2O 2.3 + 0.46

PES BSA+NaCl 30 + 19.04

PES BSA+H2O 11 + 5.13

Membrane ζ-potential (mV) - pH 4.7

ζ -potential (mV) - pH 10.0

CRC 8.6 + 0.83 9.2 + 8.15

PES 16 + 1.46 23 + 6.48

Our results for electrostatic potential test and calculated zeta potential values of protein coated membranes showed that varying pH conditions indeed affect the ionic binding and the electrostatic potential distribution. Future agenda of this research will involve modifying and integrating the streaming potential device and methods in order to test zeta potential values of cultured intestinal mucosal epithelia.

NSFDr. V.G.J. Rodgers

Dr. D.Lo B2K Group, especially Devin McBride

BRITE REU 2009 Personnel

[1] [1]

[1] Wang, Yiheng, Thesis, 2008[2] Nystrom et al. E.Streaming Potential as a tool in the characterization of ultrafiltration membranes. Colloids

and Surfaces 1989, 36, 297-312