Transcript Background: Results

Background:
Biofilms are complicated structures used
by pathogens to survive in vivo. The
immunocompetent host cannot eliminate
the pathogens in biofilms. Such
polymicrobial infections are highly
difficult to treat due to the resistance
offered by the bacteria thriving in a
symbiotic relationship enabling suitable
growth conditions for their survival. In an
attempt to recognize the organisms that
are found in biofilms, we have established
a SPR-based method to distinguish
pathogens by their interaction to specific
antibodies that were developed in vitro.
Methods
 Blood cells were separated by gradient
centrifugation.
 B-cells were cultured with and without
bacteria in L-15 cell culture medium and
incubated at 37ºC.
 Blood cell components were identified
by Flow Cytometry.
 Antibodies were produced in vitro and
were analysed by SDS-PAGE.
•Immobilization of Antibodies on CM5
sensor chip of Biacore 1000.
Bands were obtained at 25 and 50KDa
for light and heavy chains of antibodies
respectively.
Results :
Aim
The study aims to use SPR system to
detect the most common bacteria found
in biofilms to treat infections.
The medium contained antibodies
against incubated bacteria that was
immobilised on chip in Biacore system.
B cells that were incubated with
bacteria were developed to mature B
cells(CD19+ly) as shown by flow
cytometry analysis.
Ulcer secretion from healthy/patients
were analysed by Biacore and the
results correlated well to the culture
results.
Conclusion
In the present study an In vitro
model of antibody production
against bacteria is presented.
Detection of bacteria by specific
antibody interaction in SPR
system might be used to identify
the bacterial flora in secretions
especially when they are infected
by several bacterial strains or
when the culture and PCR
analyses are negative .
Detection of bacterial flora
in biological secretions
using antibodies developed
in vitro and immobilised in
a surface plasmon
resonance system
Contact Info:
Nakka Sravya Sowdamini
[email protected]
Acknowledgements
I would like to thank my
supervisor and Johanna Lonn
for their guidance throughout
the study. I am also grateful
to peas Institut for financially
supporting my study.
Master Thesis by Nakka Sravya
Sowdamini
International Masters Program, 2011
Molecular Genetics and Physiology
Supervisor: Dr Fariba Nayeri