Preparing a Hemolytic Staphylococcus aureus Culture for Host-Pathogen Interaction Studies

JoVE Encyclopedia of Experiments

Microbiologie

0 weergaven • 2:43 min • February 2nd, 2026

$$\rightleftharpoonup{xx}$$ $$\longleftharp{xx}$$, $$\longrightharp{xx}$$,

Begin with a selective nutrient agar plate supplemented with blood containing RBCs.

Streak Staphylococcus aureus (S.aureus), a Gram-positive pathogenic bacterium, onto the agar and incubate.

During incubation, bacteria utilize nutrients, grow, and form colonies.

S. aureus secretes hemolysin, a pore-forming toxin that lyses the RBCs in the agar and produces clear, transparent zones around colonies, indicative of hemolysis.

Select a hemolytic colony, inoculate it into a nutrient broth, and incubate under rotating conditions to support bacterial growth.

Using a spectrophotometer, measure optical density (OD) to determine bacterial concentration.

Based on the OD reading, transfer an aliquot into a fresh nutrient broth to dilute the cells, and incubate in a water bath with shaking.

Measure the OD at regular intervals.

An OD near 0.8 indicates that the bacteria have reached the exponential growth phase with uniform physiology.

This hemolytic S. aureus is ready for host-pathogen interaction studies.

Begin by streaking S.aureus strains of interest on blood agar plates from a frozen glycerol stock and incubating the plates at 37 degrees Celsius for 16 to 24 hours to confirm their hemolytic phenotypes based on their ability to lyse red blood cells and to form clear transparent zones.

Inoculate single colony isolates of each strain in 4 milliliters of tryptic soy broth, or TSB, in sterile glass tubes and rotate the tubes at 37 degrees Celsius for 16 to 24 hours in a tube roller at an approximately 70-degree angle and 70 rotations per minute. The next morning, measure the optical density at 600 nanometers, or OD600, of the cultures on a spectrophotometer and dilute the cells to an OD600 of 0.05 in 25 milliliters of sterile TSB at a 5 to 1 flask to volume ratio in 125 milliliter DeLong flasks. Incubate the cultures in a 37 degree Celsius water bath at 280 rotations per minute.

10:50

Kwalitatieve en Kwantitatieve Testen voor detectie en karakterisering van proteïne Antimicrobials

Gerelateerde video's

0 Bekeken

12:27

Kwantificeren van de cytotoxiciteit van Staphyloccus aureus tegen menselijke polymorfonucleaire leukocyten

Gerelateerde video's

0 Bekeken

08:32

Ontwikkeling en beoordeling van intracellulaire infectiemodellen voor Staphylococcus aureus

Gerelateerde video's

0 Bekeken

02:14

Preparation of Streptococcus mitis Culture for Host-Pathogen Interaction Studies

Gerelateerde video's

0 Bekeken

03:30

An In Vitro Assay to Study Microbial Interactions in Co-Culture

Gerelateerde video's

0 Bekeken

03:35

In Vitro Biofilm Synthesis by Staphylococcus aureus

Gerelateerde video's

0 Bekeken

03:35

Cytotoxicity Assay of Staphylococcus aureus Strains on Neutrophils following Phagocytosis

Gerelateerde video's

0 Bekeken

05:28

A Bacterial Infection Assay for Studying Anaerobic Bacteria-Host Cell Interactions

Gerelateerde video's

0 Bekeken

06:37

Staphylococcus aureus Growth met humaan hemoglobine als een ijzerbron

Gerelateerde video's

0 Bekeken

08:03

Een vloeistofchromatografie tandem-massaspectrometrie gebaseerde benadering voor metabolietanalyse van Staphylococcus aureus

Gerelateerde video's

0 Bekeken

Laatst bijgewerkt: 15 augustus 2026