Phage Titering Assay

Phage therapy is a good alternative to antibiotics by using bacteriophages to treat various infections caused by bacteria. Equipped with world-leading technology platforms and professional scientific staff, Creative Biolabs provides providing novel phage titering assays for our clients all over the world.

Introduction of Phage Titering

Bacteriophages are viruses capable of infecting bacteria. It is nucleic acid surrounded by a protein coating. Till now, a variety of bacteriophages have been extensively characterized. During later stages of phage development, different phages release progeny phage particles from infected bacteria in different ways. At the same time, infected cells are killed. When a lawn of susceptible bacterial cells is plated with bacteriophages, a circular turbidity, called a plaque, will form. Assuming that each patch is due to one phage infecting one cell, the concentration of viral particles in the stock can be determined by identifying individual patches on bacterial lawns.

A plaque-forming unit (PFU) is a measure to describe the number of phages capable of forming plaques per unit volume. Phage titering, also known as plaque assay, is the process of determining phage concentration by dilution and plating with susceptible cells. In general, lysates with a final concentration greater than 10⁹ PFU/ml are referred to as "high titer" lysates, and the higher the titer, the more stable the lysate.

Phage Titering Methods

Commonly, there are two methods for phage titering, which include whole plate titer and spot titer. Of the two, spot titer is quick and easy, while whole plate titer gives the most accurate results. To achieve this experiment, exponentially growing bacteria are exposed to phages of interest, which are then allowed to multiply overnight. Serial dilution of phage samples is the first step in both approaches and using a 10-fold dilution is the most common and easiest way. During this process, the phage is diluted enough to produce countable plaques in the bacterial lawn. Subsequently, bacteria were used for infection in each phage dilution group and cultured overnight. The next day, many plaques should be visible, and the number of plaques correlates with the respective phage dilution. At this point, the titer of the phage sample can be calculated from the plate with countable plaques. Also, the number of plaques should be reduced tenfold in samples from adjacent dilution groups.

Fig.1 Spot test and double agar overlay plaque assays.¹

Creative Biolabs offers the most advanced and comprehensive phage titering assays. We work to amplify your success in a highly collaborative manner. If you are interested in our services, please do not hesitate to contact us for detailed information.

Reference:

1. Daubie, Valéry, et al. "Determination of phage susceptibility as a clinical diagnostic tool: A routine perspective." Frontiers in Cellular and Infection Microbiology 12 (2022): 1000721. Under Open Access license CC BY 4.0, without modification.