Shigella Bactericidal Assay
Shigella Bactericidal Assay - Related Videos
06:11
A High-throughput Shigella-specific Bactericidal Assay
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Cited by 6 •
2019
Here we present a protocol to measure Shigellacidal activity of antibodies in serum. Serum is mixed with bacteria and exogenous complement, incubated, and the reaction mixture is plated on agar plates. Viable bacteria form colonies which are counted, using an automated colony enumerator, and used to determine the bactericidal titer.
05:40
A Serum Bactericidal Assay for the Complement-Mediated Bactericidal Activity of Antibodies
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2025
This video demonstrates a technique to evaluate the complement-mediated bactericidal activity of antibodies present in serum. This approach combines isolated serum with pathogenic bacteria and exogenous complement proteins. Upon incubation, the bacteria are grown on an agar plate, and the produced colonies are quantified to measure the serum bactericidal titer.
04:22
Intracellular Replication Assay for Shigella in Human Colonic Epithelial Cells
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2026
Source: Poore, K., et al. Epithelial Cell Infection Analyses with Shigella. J. Vis. Exp. (2024)This video demonstrates an in vitro assay to assess Shigella invasion and intracellular replication in human colonic epithelial monolayers. It highlights steps for infection, antibiotic selection, host cell lysis, and recovery of intracellular bacteria for downstream analysis.
02:51
Stress Granule Formation During Shigella Infection of Cancerous Epithelial Cells
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2026
Source: Vonaesch, P. et. al., Immunofluorescence Analysis of Stress Granule Formation After Bacterial Challenge of Mammalian Cells. J. Vis. Exp. (2017)This video demonstrates the infection of epithelial cells with the pathogenic bacterium Shigella flexneri and the resulting host cellular response. Bacterial effector proteins disrupt host translation, triggering the formation of membrane-less organelles called stress granules, which reflect the cell's adaptive response to infection.
02:55
Determining the Role of Vitronectin in Bacterial Resistance to Bactericidal Activity of Human Serum
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2025
This video demonstrates the role of vitronectin, a human serum glycoprotein, on bacterial resistance. Antibodies in the serum interact with bacterial surface proteins, activating complement proteins and forming the membrane attack complex, causing bacterial cell lysis. Vitronectin binding on bacterial membrane protein prevents the membrane attack complex formation, resulting in bacterial survival.
FAQs
Antibodies first recognize and bind surface antigens on Shigella cells. This binding enables complement activation under the assay conditions, linking antigen recognition to a killing response. The measured activity therefore reflects cooperation between serum antibodies and complement rather than antibody attachment alone, making the result relevant to functional antibacterial immunity.
Binding assays indicate whether antibodies recognize Shigella antigens, but they do not necessarily show whether those antibodies produce a biological effect. The bactericidal assay adds this functional dimension by measuring the reduction in viable bacteria after serum exposure. Consequently, two sera with detectable binding antibodies may display different levels of antibacterial activity.
Defined conditions are important because antibody binding, complement activation, and bacterial survival must be assessed in a consistent setting. When test sera are evaluated under the same conditions and compared with appropriate controls, differences in surviving Shigella can be interpreted as differences in functional serum activity. This standardization supports comparisons among samples.
The workflow begins by combining Shigella cells with a test serum under defined assay conditions. The mixture is incubated so antibodies can bind bacterial surface antigens and activate complement. Researchers then quantify surviving bacteria by culture and compare that measurement with controls, producing a functional readout of serum-associated antibacterial activity.
Controls establish the comparison needed to determine how much bacterial survival changes in the presence of the test serum. Culture-based quantification supplies the experimental outcome by measuring viable Shigella after incubation. Together, these elements help distinguish serum-associated killing from the baseline bacterial survival represented by the assay controls.
Researchers apply the assay to evaluate immune responses generated by Shigella vaccine candidates, compare the functional activity of serum samples, and investigate responses relevant to protection against infection. Because it measures antibacterial action rather than antibody presence alone, the method can also support efforts to identify immunological correlates of protection.