OWLS APPLICATION NOTES NO-001

LABEL-FREE IMMUNOSENSOR
FOR HSP70 DETECTION
using OPTICAL WAVEGUIDE LIGHTMODE SPECTROSCOPY (OWLS) detection
Abstract
OWLS immunosensor offers a real time label-free detection of heat shock protein, HSP70 (molecular weight of
70 kDA). For biosensing monoclonal antibodies raised against HSP70 protein under investigation were used. To
form regenerable sensor surface, the waveguide sensor surface was modified with amino group and sensitized by
immobilizing antibody/antigen molecules covalently to the surface.
Application of OWLS sensors
as immunosensor for the detection of the heat- Calibration curve for HSP-70 sensor
shock protein HSP70

12

10

Se nsor re sponse (a.u.)

HSP70
sample
monoclonal antibody 8

100 - 150 nm
6
laser light waveguide
4

EM field glass
distribution support 2
0.0001 0.001 0.01 0.1 1 10 100

HSP70 conce ntration (ng/ml)

Surface Chemistry
• Amino functionalisation of waveguide surface Measurement of anti-HSP70 antibody
by (γ-aminopropyl)triethoxysilane HSP70 molecules are covalently attached to the
• Immobilisation of biomolecules on the amino surface and different dilutions of anti-HSP70
surface of the OW LS sensors by a n t i b o d y we r e m e a s u r e d .
glutaraldehyde.
20 µ g/ml
Sensitization of the sensor surface 400

with monochlonal anti-HSP70 IgG 10 µ g/ml

350
Sensor response (a.u.)

5 µ g/ml

600 300 1 µ g/ml

HCl HCl

500
250
Sensor response (a.u.)

400
Blocking
200
300

MoAb
200
150
GA
100 70 90 110 130 150
TRIS Time (min)
0
0 20 40 60 80 100 120 140
Time (min)

Conclusion
Regeneration cycles of the measurement The OW LS Sensor is a very sensiti ve and cost-
Antibodies are covalently attached to the surface effective tool for HSP70 and anti-HSP70 antibody
a n d l i f e t i m e o f s e n s o r s we r e e x a m i n e d a n d detection.
optimized. The lowest detectable amount of HSP70 is 1 pg/ml.

500
References
10 pg/ml 20 pg/ml
5 pg/ml
1 pg/ml 1. Vörös, J. J. Ramsden, G. Csucs, I. Szendrõ,
450 S . M . D e P a u l , M . T e xt o r , N . D . S p e n c e r
Sensor response (a.u.) xx

“Optical Grating Coupler Biosensors”

400 Biomaterials 23 (2002) 3699-3710

350 2. Levkovets I., Adányi N., Trummer N., Váradi

M., Szendrı I., Starodub N.F., Székács A.,
300 “Developm ent of optical (O W LS)
immunosensors for macromolecules and
250
s m a l l a n a l y t e s ” , B I O K É M I A X XV I I I / 1 , 2 0 0 4
100 150 200 250 300
Time (min) 3. w w w . o wl s - s e n s o r s . c o m

MicroVacuum Ltd. H-1147, Kerékgyártó u. 10., Budapest, Hungary Phone: + 36 1 252 1991; +36 1 467 0108 Fax: +36 1 221 7996
Web: http://www.owls-sensors.com/; E-mail: info@owls-sensors.com Copyright @ Microvacuum, 2006