Fluorescent Probes for Imaging Bacteria

Fluorescent probes for imaging bacteria are useful for advancing bacterial research and antibiotic design. One technique for studying bacteria, is to incorporate fluorescent probes such as Fluorescent D-amino acids (FDAAs) into the cell wall of live bacteria. FDAAs are suitable for use with confocal and super-resolution microscopy (SRM).

Literature (1)
Cat. No. Product Name / Activity
6798 Click N-Acetylmuramic acid - alkyne
Bacterial peptidoglycan derivative; suitable for 'click'-conjugation to fluorescent dyes
7506 Click N-Acetylmuramic acid - azide
Bacterial peptidoglycan derivative; suitable for 'click'-conjugation to fluorescent dyes
7714 EDA-DA
Unnatural dipeptide building block with alkyne group for functionalizing peptidoglycan
6647 HADA
Blue fluorescent D-amino acid for labeling peptidoglycans in live bacteria
6648 NADA-green
Fluorescent D-amino acid for labeling peptidoglycans in live bacteria
7408 OGDA
Green fluorescent D-amino acid; compatible with STED microscopy
6649 RADA
Orange-red TAMRA-based fluorescent D-amino acid for labeling peptidoglycans in live bacteria
7406 Rf470DL
Blue rotor-fluorogenic fluorescent D-amino acid for labeling peptidoglycans in live bacteria
7860 sBADA
Green fluorescent D-amino acid for labeling peptidoglycans in bacteria
7449 Se-NADA
Orange fluorescent benzoselenadiazole D-amino acid (FDAA) for imaging bacteria; also photosensitizer
6802 6 TMR Tre
Fluorescent trehalose; selectively labels mycobacterial cell envelope
6650 YADA
Green-yellow lucifer yellow-based fluorescent D-amino acid for labeling peptidoglycans in live bacteria

Antibiotic Research: Targeting peptidoglycans (PG)

The investigation of bacterial physiology is essential for developing antibiotics to treat diseases such as E. coli infection. Targeting PG biosynthesis is a common approach to designing antibiotics, since it is essential for bacterial growth and function, as well as being selectively lethal to bacteria without harming human cells.

PGs are components of bacterial cell wall, which protect the bacteria from environmental stresses. In most bacteria, a peptidoglycan cell wall is made up of macromolecular polymers of glycan strands that link to the cytoplasmic membrane via oligopeptide bridges.

Bacteria are split into two groups, gram-negative and gram-positive, depending on the organization of the PGs. Gram-negative bacteria have a PG layer of about 5 nm, between a cytoplasmic membrane and the outer membrane; gram-positive bacteria have a much thicker PG layer of about 20-50 nm on the periplasmic side of the cytoplasmic membrane. Mycobacteria do not fit in to either category, they have a PG layer and a waxy surface layer composed of mycolic acids, lipids and arabinogalactan.

The ability to investigate PG physiology in high resolution is a useful development for bacterial research and antibiotic design. New SRM techniques are enabling researchers to study bacteria in high resolution under physiologically relevant conditions to identify new therapeutic vulnerabilities.

Techniques for Imaging and Staining Bacteria

Previous techniques used for observing bacteria include electron microscopy, but this method had the disadvantage of using mercury-containing chemicals. The fixation and staining required for this technique precluded work with live bacteria. The next advancements came from fluorescent microscopy which has been useful for gaining a greater understanding of bacterial physiology and function in live cells. However, this technique is limited because spatial resolution is restricted by the diffraction limit of incident light to ~250 nm (Abbe diffraction limit). Recent advances of both super-resolution microscopy techniques (including STORM, SIM, STED and PALM) and new molecular probe types (namely Fluorescent D-amino acids (FDAAs)) has accelerated the understanding of peptidoglycan biosynthesis dynamics and mechanisms, allowing unprecedented analysis at a molecular level.

Using FDAAs to Stain Bacteria

Previously fluorescently-labeled antibiotics, and fluorescently-labeled wheat germ agglutinin (FWGA) were used as molecular probes to study PGs. Both of these probes have major drawbacks, fluorescent antibiotics suppress bacterial growth, and FWGA cannot permeate the cell walls of gram-negative bacteria.

FDAAs are metabolic probes that leverage the promiscuous nature of the bacterial PG 60 biosynthetic pathway. FDAAs are small molecules conjugated to D-amino acids and are incorporated into new poly-glycan chains. They efficiently label peptidoglycans in bacterial cell walls in situ, allowing the study of bacteria morphology and formation, as well as bacterial growth. FDAAs can be used in gram-negative and gram-positive bacterial species, in real-time and in live cells, with minimal toxic effects. They are suitable for use with both confocal and super-resolution microscopy.

Spectral graphs for the FDAA range

Figure 1: The Tocris Fluorescent D-amino acids (FDAAs) collection spans the visible spectrum, giving useful multiplexing options. For more details read Hsu et al (2017) Full color palette of fluorescent d-amino acids for in situ labeling of bacterial cell walls. Chem.Sci. 8 6313 PMID: 28989665

5 star FDAA review - featuring HADA (Cat. No. 6647)

hada image

See our 5 star HADA review, written by Dr Patrick Moynihan. This review shows HADA labeling peptidoglycan in growing mycobacterial species.

Customers can leave reviews and share their images for all our products, as well as leaving tips and details of protocols that helped with their research. This helps the research community build up confidence in good products and protocols and as a thank you from Tocris for your time, we give out amazon gift cards.

Literature for Fluorescent Probes for Imaging Bacteria

Tocris offers the following scientific literature for Fluorescent Probes for Imaging Bacteria to showcase our products. We invite you to request* your copy today!

*Please note that Tocris will only send literature to established scientific business / institute addresses.

Fluorescent Dyes and Probes Research Product Guide

Fluorescent Dyes and Probes Research Product Guide

This product guide provides a background to the use of Fluorescent Dyes and Probes, as well as a comprehensive list of our:

  • Fluorescent Dyes, including dyes for flow cytometry
  • Fluorescent Probes and Stains, including our new MitoBrilliantTM mitochondria stains
  • Tissue Clearing Kits and Reagents
  • Aptamer-based RNA Imaging Reagents
  • Fluorescent Probes for Imaging Bacteria
  • TSA VividTM Fluorophore Kits
  • TSA Reagents for Enhancing IHC, ICC & FISH Signals