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Super Resolution Microscopy Imaging Tools – high quality, low cost

Janelia Fluor® Preadsorbed Antibodies | Janelia Fluor® Conjugation Kits | Streptavidin Conjugates | Dyes: Amine-Reactive, Click-Active, Free-Acid

What is Super Resolution Microscopy?

  • Super resolution microscopy (SRM) techniques enable the resolving of structures that are beyond the diffraction-limit resolution of traditional light microscopy (~200nm), allowing production of high resolution images.

  • Various SRM techniques including stimulated emission depletion (STED) microscopy, structured illumination microscopy (SIM), direct stochastic optical reconstruction microscopy (dSTORM) and photoactivation localization microscopy (PALM).

  • For optimal results the specific probe or dye used for each technique will vary, as each SRM technique has specific fluorophore requirements. For example, the dSTORM technique is based on use of organic dyes such as Janelia Fluor® as photoswitchers.

  • Some SRM techniques also require specific buffers to help regulate the photophysical properties of fluorophores used.

                                                   

                                        Multicolor STED image of a COS-7 cell. Adapted from [1]

Advantages of Janelia Fluor® dyes

  • Janelia Fluor® dyes are exceptionally bright, photostable and cell permeable dyes suitable for SRM techniques such as dSTORM and STED as well as live-cell imaging, confocal microscopy, immunocytochemistry and HaloTag® and SNAP-tag® systems.
  • Janelia Fluor® dyes are novel fluorescent dyes which are derived from the classic fluorophore tetramethylrhodamine (TMR) with the N,N-dimethyamino groups replaced with azetidine substitutes; a novel, simple modification in the fluorophore structure enabling improved brightness without negatively impacting molecular size and cell permeability.
  • Janelia Fluor® conjugates outperform other fluorophores in terms of signal intensity and demonstrate higher resistance to photobleaching. 

             

Figure 1. Absorption and emission spectra of Janelia Fluor® dyes. Adapted from Janelia Research Campus [2]. 

Figure 2. Superior anti-fade performance of Janelia Fluor® 549 conjugated dyes compared to those conjugated with DyLight 550 fluorophores. 

 

Figure 3. MBP staining in rat cerebellum using Goat Anti-Mouse IgG H&L Janelia Fluor® 549 (HB9240) preadsorbed secondary antibody.

 

Super Resolution Imaging Tools Available from Hello Bio

Hello Bio offer a range of high quality yet affordable imaging tools for use in Super Resolution Microscopy (SRM) including: 

Secondary antibodies

Discover our range of Janelia Fluor® preadsorbed antibodies as part of ValidAb™ range of highly validated, data-rich antibodies.   
                        

                     

Rat brain stained for GFAP using Goat Anti-Mouse IgG H&L Janelia Fluor® 525 (HB8144) preadsorbed secondary antibody 

 

Conjugation kits

We offer the novel Janelia Fluor® conjugation kits for labeling antibodies and proteins with Janelia Fluor® 525 and Janelia Fluor® 549 in as little as 90 minutes.

                                         Rat hippocampus stained with Janelia Fluor 549<sup>®</sup> conjugated anti-NeuN antibody

     Rat hippocampus stained with Janelia Fluor® 549 conjugated anti-NeuN antibody

Streptavidin

Streptavidin-based amplification techniques allow for improved detection sensitivity and boosts signal from low-abundance targets. Hello Bio offers a variety of streptavidin conjugates for detection and signal amplification of biotin coupled proteins and antibodies.

                 

Rat hippocampus stained with parvalbumin anti-mouse biotin antibody followed by incubation with Streptavidin Janelia Fluor® 525 (HB15382)

 

Tyramide Signal Amplification (TSA)

Tyramide signal amplification (TSA) utilizes horse radish peroxidase (HRP) to enzymatically convert fluorophore tyramides to bind tyrosine residues on and surrounding the protein epitope targeted by the primary antibody.

***COMING SOON!***

                     

Rat hippocampus stained for parvalbumin anti-mouse ValidAbTM antibody detected using Biotin TSA Kit - AF488 (HB18560) compared to conjugation with streptavidin AF488 (HB13531).

Dyes

Hello Bio offer a variety of high quality yet affordable fluorescent dyes to label biomolecules for visualization by fluorescence microscopy, in a range of reactive groups including:

  • amine-reactive supplied as an NHS ester for coupling to primary amine (NHS) groups
  • click-active supplied as azide or tetrazine for use in click-chemistry.
  • free acid used for synthesis of Janelia Fluor® HaloTag® and SNAP-Tag® ligands for use in live cell imaging experiments. Also suitable for flow cytometry.

         

Excitation and emission spectra of Janelia Fluor® 549, free acid (HB8745)

 

Protocols & Resources:

We have a detailed and comprehensive range of protocols and resources to support you:

Protocols

Methods & Guides

Further Reading:

[1] Image adapted from Dr. Eugene Katrukha, Utrecht University, The Netherlands.

[2] Jacquemet, G. et al. (2020). The cell biologist's guide to super-resolution microscopy. J Cell Sci 1 June 2020; 133 (11): jcs240713.

[3] Janelia Fluor® dyes: https://www.janelia.org/open-science/janelia-fluor-dyes

[4] Grimm JB, et al. (2017). Synthesis of Janelia Fluor HaloTag and SNAP-Tag Ligands and Their Use in Cellular Imaging Experiments. Methods Mol Biol. 2017;1663:179-188. doi: 10.1007/978-1-4939-7265-4_15.

[5] Lelek M. et al. (2021). Single-molecule localization microscopy. Nat Rev Methods Primers. 2021;1:39. doi: 10.1038/s43586-021-00038-x.