Intracellular Bacteria Isolation
Intracellular Bacteria Isolation - Related Videos
03:41
Isolation of Intracellular Pathogenic Bacteria from Infected Host Cells
0 Views •
2026
Source: Sigal, N., et al. RNA Purification from Intracellularly Grown Listeria monocytogenes in Macrophage Cells. J. Vis. Exp. (2016).This video demonstrates the isolation of internalized pathogenic bacteria from infected bone marrow-derived macrophages (BMDMs). Bacteria are allowed to attach to host cells, and unattached bacteria are washed away. After internalization, an antibiotic is added to eliminate non-internalized bacteria. The host cells are lysed, centrifuged, and the supernatant is...
01:33
Investigating Autophagy-Mediated Clearance of Intracellular Bacteria In Vitro
0 Views •
2026
Take pig kidney cell cultures, with one culture pre-treated with an inhibitor that increases lysosomal pH, impairing autophagic-lysosomal degradation.Add pathogenic bacteria and centrifuge to enhance host-pathogen contact.Incubate to allow bacterial invasion and internalization into endocytic vesicles.The internalized bacteria consume host nutrients to replicate, triggering metabolic stress.This stress activates autophagy, where an autophagic membrane expands around the vesicle, forming an...
01:10
Intracellular Movement of Viruses and Bacteria
0 Views •
2023
Intracellular bacteria and viruses often comprise a group of highly infectious pathogens that can cause several diseases. Bacterial pathogens include those belonging to the genus Rickettsia responsible for conditions such as rocky mountain spotted fever and the Mediterranean spotted fever; Chlamydia, a genus responsible for a sexually transmitted disease; Coxiella burnetii, an agent responsible for Q fever. Viral pathogens include vaccinia—a poxvirus, and herpes simplex virus—a virus that...
02:40
Isolation of Bacteria from the Mosquito Midgut
0 Views •
2026
Source: Liu, X., et al. A Bacterial Oral Feeding Assay with Antibiotic-Treated Mosquitoes. J. Vis. Exp. (2020)This video demonstrates the dissection of the mosquito midgut to isolate gut bacteria. The midgut is extracted from the digestive tract, homogenized to release the bacteria, and plated on nutrient agar to culture bacterial colonies.
02:56
Isolation of Mouse Bladder Epithelial Cells Containing Intracellular Bacterial Communities
0 Views •
2026
Source: Yang, E., et al. Isolation of Single Intracellular Bacterial Communities Generated from a Murine Model of Urinary Tract Infection for Downstream Single-cell Analysis. J. Vis. Exp. (2019)This video demonstrates the manual isolation of single epithelial cells containing intracellular bacterial communities from infected mouse bladder using mouth micropipetting, enabling downstream single-cell analysis of viable host-associated E. coli populations in urinary tract infection...
FAQs
Selective host-cell lysis breaks open eukaryotic cells while aiming to preserve the bacterial cells released from them. This creates a critical separation step: excessive disruption may damage bacterial viability, whereas insufficient lysis can leave bacteria trapped within host material. Balancing these conditions improves recovery and supports later culture, counting, microscopy, or molecular analysis.
Controlled washing reduces bacteria present outside the host cells before lysis begins. This distinction matters because extracellular microbes can be mistakenly counted or analyzed as intracellular populations, obscuring host-associated behavior. Washing therefore improves interpretive accuracy, although the process must remain controlled so that host cells and their internal bacterial contents are not unnecessarily lost.
After host-cell disruption, differential centrifugation or density-based separation helps partition bacterial cells from host-cell material according to their physical separation behavior. These approaches provide complementary ways to enrich the bacterial fraction while reducing cellular debris. The resulting preparation is more suitable for downstream culture, enumeration, characterization, microscopy, or molecular assays than an unseparated lysate.
The balance among washing, host-cell lysis, and separation conditions largely determines recovery, viability, and contamination. Conditions that are too harsh can compromise bacterial survival, while conditions that are too mild may leave host material or extracellular microbes in the preparation. Adjusting these steps helps produce material that more accurately represents bacteria residing within host cells.
A typical workflow begins by washing the host-cell preparation to reduce extracellular microbes, followed by controlled lysis of the eukaryotic cells. The released material is then processed using differential centrifugation or density-based separation to reduce host-cell components. The recovered bacterial fraction can subsequently be cultured, counted, characterized, or examined through microscopy and molecular assays.
Researchers use the recovered bacterial material when they need to examine behavior associated with life inside host cells rather than in the surrounding environment. The material supports studies of host–pathogen interactions, intracellular survival, and antibiotic susceptibility. It can also provide samples for characterization and molecular analysis, helping connect bacterial recovery with mechanisms of infection.