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http://hdl.handle.net/11375/26657
Title: | Detection and quantification of Legionella pneumophila using latex agglutination test |
Authors: | Mathai, Aiswarya |
Advisor: | Puri, Ishwar |
Department: | Biomedical Engineering |
Publication Date: | 2021 |
Abstract: | Legionella pneumophila (L. pneumophila) are ubiquitous in environmental water sources that are responsible for causing sporadic and epidemic cases of atypical pneumonia, called Legionnaires’ disease (LD), after inhalation of contaminated water droplets. There is a need for a rapid detection method for Legionella pneumophila fluid samples without the need for complex instrumentation. Agglutination assays are widely used in biology and medicine for their simplicity and specificity. However, conventional agglutination tests only provide presumptive results. Here I describe and perform a latex agglutination test (LAT) that can detect and quantify L. pneumophila in a sample by determining the degree of agglutination. The latex microbeads (beads) are coated with polyclonal antibodies using established methods. The optimal ratio of the beads to the antibody for the best performance of LAT was found to be 20:3. L. pneumophila serogroup 1 is chosen as the target bacteria and Escherichia coli K12 as the negative control. Strains of L. pneumophila serogroup 1 were cultured and inactivated by heat treatment for safety. The target bacteria displaying the antigenic epitopes were mixed with the antibody-coated beads and have been shown to facilitate agglutination. Micro-channels fabricated using adhesive tapes are used to entrap the fluid sample, where an image analysis algorithm quantifies the degree of agglutination using images obtained by a microscope, thereby, resolving issues with complex techniques. Specific binding between antibody and L. pneumophila serogroup 1 is investigated by tracking the motion of the beads and bacteria in the images. The number of beads involved in agglutination compared to the total number of beads present in an image determines the degree of agglutination. The sample volume used for detection is 3 µL and the detection limit of the method is 106 cells/ml. Compared to expensive and complex PCR techniques and time-consuming culture methods, this LAT is simple, affordable, and can produce results within hours using only a microscope. |
URI: | http://hdl.handle.net/11375/26657 |
Appears in Collections: | Open Access Dissertations and Theses |
Files in This Item:
File | Description | Size | Format | |
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Mathai_Aiswarya_S_2021June_Master of Applied Science.pdf | 1.64 MB | Adobe PDF | View/Open |
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