1. Using an Enzyme-Linked Immunosorbent Assay (ELISA) for the Detection of
Streptococcus pneumoniae in Human Bodily Fluids
Jermaine D. Dorsey and Mamie T. Coats
Department of Biological Sciences, Alabama State University, Montgomery Alabama, USA, 36101
Abstract
Streptococcus pneumoniae is a major cause of
bacteremia, meningitis, and pneumonia in both
children and adults. Invasive pneumococcal
disease (IPD) primarily affects young children, older
adults (> 65 years of age), and individuals with
comorbidities or impaired immune systems. The
diagnosis of pneumococcal infections has been
attempted using methods which detect specific
antigens by the procedures of a quantitative
enzyme-linked immunosorbent assay (ELISA). We
will detect the antigen D39, WU2, and EF3030 for
the diagnosis of Streptococcus pneumoniae
infection.
Introduction
Streptococcus pneumoniae, pneumococcus, is a
gram-positive pathogen found in the upper respiratory
tract of healthy children and adults. Worldwide, S.
pneumoniae remains one of the leading cause of
community acquired bacterial infection for diseases
like; pneumonia, otitis media, bacterial meningitis,
and bacterium.
Pneumococcus polysaccharide capsule surrounds
the pneumococcus and protects it against clearance
by phagocytosis. More than 90 different capsule
serotypes have been discovered thus far.
When pneumococcal infection is suspected time is
very important in influencing patient outcomes.
Classical identification takes several days to verify
and molecular techniques are limited due to the
variability of the capsular polysaccharide. The overall
goal for this research is to develop a rapid, serotype
independent assay to detect S. pneumoniae in body
fluids in using an Enzyme-Linked Immuno-Sorbant
Assay.
.
• Coated 96-well plate with PspC specific polyclonal capture
antibody
• Blocked with Bovine Serum Albumin (BSA)
• Added 10X5 CFU/mL S. pneumoniae (strain D39, WU2)
• Added Detection antibody with incubation at 37°C
• Added labeled 2 antibody
• Added Substrate
• Read OD405
Discussion and future plan
• Overall, our experiment has shown that we
can detect PspC through ELISA Method.
• Use the same method in presence of bodily
fluids to determine the sensitivity of this
method of detection.
• Other virulence factors will be examined as
potential capture antigens.
Acknowledgements
I would like to thank my mentor and MARC U
STAR program, Biomedical Research and
Training Program at Alabama State University,
Montgomery, Al. 36104. This work was
supported by the National Institutes of Health
(NIH).
Methods and Results
Capture Antibody
Target Antigen
Detection
Antibody
Labeled Antibody
Biotin
Substrate
Anticipated results
0
1
2
3
4
5
6
7
1 2 3 4 5 6 7 8 9
LevelofPspCinEnvironment
(media)
Log CFU/mL
Free PspC
The level of
PspC in the
growth
environment is
directly
proportional to
the number of
pneumococci
present
The level of PspC
in the growth
patients’ samples
is directly
proportional to the
level of infection
and the site of
infection
pathmicro.med.sc.edu
0
2
4
6
8
10
12
Sputum
Blood
Infection level
AmountofPspCdetected