Protein Adsorption on Solid Surfaces

Chemical Engineering and Materials

Kim-Lien Thi DinhMajor: Biochemistry

Pasadena City College

Mentor: Kenny RosenbergFaculty Advisor: Jacob Israelachvili

University of California, Santa Barbara

• Fundamental: Understand the interactions of protein– Protein-protein interactions– Surface-protein interactions

• How much protein is on the surface?• What would happen if…?

• Extend the life span of a biomedical implant– Implant rejected by the body due to protein adsorption– Decrease the adsorption of protein

• Funding: National Institute of Health– Improve people’s health– Prevent diseases/find causes and cures for diseases

Research and Motivation

Surface-Protein Interactions

• Proteins– BSA (bovine serum albumin)– Tau

• Need a “balance”– Quartz Crystal Microbalance (QCM)

• Super-sensitive balance• Detects the mass of adsorbed protein

• Surfaces– Gold– Mica

• Glue it on the crystal• Make sure it works

How much protein is on the surface?

balance

0.0000 g

balance

1.0000 g

balance

0.0000 g

balance

3.0000 g

Q-Sense D300

q-sense.com/Q-Sense D300 User Manual

14,999,900.00 Hz

-100 Hz

15,000,000.00 Hz (15 MHz)

14,999,950.00 Hz

-50 Hz

15,000,000.00 Hz (15 MHz)

Sample Data5 mg/ml BSA with 200 mM NaCl (15 Mhz)

-120-100-80-60-40-20

020

0 15 30 45 60 75 90 105

Time (minutes)

∆ Fr

eque

ncy

(Hz)

How much BSA protein is on the gold surface?

24.95

25.05

25.15

0 15 30 45 60 75 90 105

Time (minutes)Tem

p(C

elci

us)

ut

-0.4

0

0.4

0.8

1.2

0 15 30 45 60 75 90 105

Time (minutes)

Dis

sipa

tion

X 1E

-6

- 80 Hz ≈ 22 ng

- 95 Hz ≈ 26 ng

Addition of BSA

Dissipation

Sum of all energy losses in the system per

oscillation cycle

Soft/rigid film

1 monolayer of BSA ≈ 15 ng

0 µm

1 µm

0.5 µm

1 µm

0.5 µm

0 µm

1 µm

0.5 µm

1 µm

0.5 µm

Gold vs. Mica

0 nm

10 nm

5 nm

0.25 µm

GOLD

AFM

0.25 µm

MICA

AFM0 nm

3 nm

2 nm

Preliminaries: 1mg/ml BSA on Mica

2.5 µm

0 µm

0 nm

2 nm

1 nm

0.5 µm

AFM

pH 10.15

2.5 µm

0 µm

Does the pH of the solution matter?

0 nm

2 nm

1 nm

0.5 µm

AFM

pH 6.80

0 µm

2.5 µm

pH 4.77

0 nm

5 nm

2.5 nm

0.5 µm

AFM

Preliminaries: Tau on MicaDoes Tau concentration matter?

0 nm

2 nm

1 nm

0 nm

2 nm

1 nm

0.25 µm

AFM

0.25 µm

AFM

1mg/ml BSA on GoldDoes the pH of the solution matter?

BSA on GoldDoes the pH of the

solution matter?

1 monolayer of BSA ≈ 15 ng

- 170 Hz ≈ 48 ng (3 layers)

- 13 Hz ≈ 3.5 ng

1mg/ml BSA with Acetic Buffer, pH 4.77

-200

-160

-120

-80

-40

0

40

0 10 20 30 40 50 60 70 80 90 100

Time (minutes)

∆ Fr

eque

ncy

(Hz)

1 mg/ml BSA with Carbonate Buffer pH 10.15

-20

-15

-10

-5

0

5

0 50 100 150 200 250

Time (minutes)

∆ Fr

eque

ncy

(Hz)

First rinse

Addition of BSA

First rinse

Addition of BSA

BSA on GoldDoes salt concentration

matter?

1 monolayer of BSA ≈ 15 ng

-80 Hz ≈ 22 ng

- 300 Hz ≈ 83 ng (5.5 layers)

1 mg/ml BSA with K-PIPES, 1000mM NaNO3, pH 6.80

-400

-300

-200

-100

0

100

0 5 10 15 20 25 30

Time (minutes)

∆ Fr

eque

ncy

(Hz)

1 mg/ml BSA with K-PIPES, 1mM NaNO3, pH 6.8

-100

-80

-60

-40

-20

0

20

0 5 10 15 20 25 30

Time (minutes)

∆ Fr

eque

ncy

(Hz)

First rinse

Second rinse

Addition of BSA

Addition of BSA First rinse

• It takes less than 5 seconds for the majority of the protein to adsorb onto the surface

• pH of the solution matters• Salt concentration matters too

• Produce a mica-modified crystal that works• Study Tau

Summary and Future Research

Acknowledgements

• INSET– Trevor Hirst– Dr. Nick Arnold– Liu-Yen Kramer– Mike Northen

• Stu Feinstein• UCSB, CNSI, PCC, ME

KENNY ROSENBERGJacob Israelachvili and friends