Zebrafish brain with green stained nuclei and orange labeled axons in the dorsonasal retina

Cell tracers are non-toxic probes that can track the flow of capillaries and define neuronal cell connectivity and translocation to monitor movement and localization in single cells, whole cell populations, or intact organisms. These fluorescent tracers can be introduced through iontophoresis, microinjection, or pinocytosis. Additionally, transport can be retrograde, anterograde, or bidirectional. While most of these fluorescent probes are effective in experiments involving fresh tissue sections and cultured cells, there are special requirements for fixed tissues and in vivo imaging.

See cell tracers selection guide

Properties of cell tracers

 Cholera toxin subunit BDiI and derivativesHydrazides and Biocytins
ReadoutDirect fluorescence imaging of cells, selective for ganglioside GM1Labels cell membranes and uniformly label neurons via lateral diffusion in the plasma membraneDirect fluorescence imaging of cells
Tissue sectionsYesYesYes
In vivo neuronsNoYesYes
Fixed tissue sectionsNoYesNo
Cultured cellsYesYesYes
Cell loadingMicroinjection, iontophoresisMicroinjection or iontophoresis; for tissue sections, crystal and paste placement options availableMicroinjection or pinocytosis
Staining modeExogenousExogenousEndogenous
Best forCell populationsCell populationsSingle cells
FixableYesNoYes
TransportRetrogradeBidirectionalBidirectional
Photobleaching resistance
Gap junction transferNoNoYes

Selection guide for cell tracers

These conjugates are useful for tissue sections, and cultured cells. They are fixable and photostable retrograde tracers introduced via microinjection or iontophoresis.

Learn more about cholera toxin subunit B

 CarrierStandard filter set(s)FluorophoreEx/Em (nm)QuantityCat. No.
Cholera toxin subunit B
Cholera Toxin Subunit B, Alexa Fluor 488 ConjugateCholera toxin subunit B (Recombinant)FITCAlexa Fluor 488495/519100 µgC34775
Cholera Toxin Subunit B, Alexa Fluor 555 ConjugateTRITCAlexa Fluor 555555/565100 µgC34776
Cholera Toxin Subunit B, Alexa Fluor 594 ConjugateTexas RedAlexa Fluor 594590/617100 µgC34777
Cholera Toxin Subunit B, Alexa Fluor 647 ConjugateCy5Alexa Fluor 647650/668100 µgC34778
Cholera Toxin Subunit B, Biotin-XX ConjugateNABiotin-XXNA100 µgC34779
Cholera Toxin Subunit B, Horseradish Peroxidase ConjugateNAHorseradish peroxidaseNA100 µgC34780
Lipid raft kits
Vybrant Alexa Fluor 488 Lipid Raft Labeling KitCholera toxin subunit B (Recombinant) and anti-cholera toxin subunit B antibody (anti-CT-B)FITCAlexa Fluor 488495/5191 kitV34403
Vybrant Alexa Fluor 555 Lipid Raft Labeling KitTRITCAlexa Fluor 555555/5651 kitV34404
Vybrant Alexa Fluor 594 Lipid Raft Labeling KitTexas RedAlexa Fluor 594590/6171 kitV34405

These cell tracers, also called lipophilic tracers, are useful for tissue sections, in vivo neurons, fixed tissue sections, and cultured cells. They are anterograde and retrograde tracers used for long term studies.

Learn more about DiI and its derivatives

 Standard filter set (s)Ex/Em (nm)FormatFixableQuantityCat. No.
DiI and derivatives
DiI Stain – DiIC18(3)Rhodamine549/565solidNo100 mgD282
DiI Stain – DiIC18(3), crystallinecrystal25 mgD3911
DiD – DiIC18(5)Cy5644/663solid10 mgD7757
DiR – DiIC18(7)Cy5.5750/780solid10 mgD12731
CellTracker DiI
CellTracker CM-DiI Dye, solutionRhodamine549/565solutionYes20 x 50 µgC7000
CellTracker CM-DiI Dyesolid1 mgC7001
Neurotrace DiI
NeuroTrace DiI Tissue-Labeling PasteRhodamine549/565pasteNo500 mgN22880
NeuroTrace CM-DiI Tissue-Labeling PastepasteYes100 mgN22883
Vybrant DiI and derivatives
Vybrant DiO Cell-Labeling SolutionGFP484/501solutionNo1 mLV22886
Vybrant DiI Cell-Labeing SolutionRhodamine549/565solutionNo1 mLV22885
Vybrant CM-DiI Cell-Labeling SolutionsolutionYes1 mLV22888
Vybrant DiD Cell-Labeling SolutionCy5644/663solutionNo1 mLV22887
Vybrant Multicolor Cell-Labeling KitVarioussolutionNo1 mL eachV22889

These cell tracers, also called polar tracers, are useful for tissue sections, in vivo neurons, and cultured cells. They are anterograde and retrograde tracers used to trace neuronal projections and investigate gap junctions.

Learn more about hydrazides and biocytins

 CarrierStandard filter set(s)FluorophoreEx/Em (nm)QuantityCat. No.
Hydrazides
Alexa Fluor 350 HydrazideHydrazide, sodium saltDAPIAlexa Fluor 350346/4425 mgA10439
Alexa Fluor 488 HydrazideFITCAlexa Fluor 488495/519A10436
Alexa Fluor 568 HydrazideRhodamineAlexa Fluor 568578/603A10437
Alexa Fluor 594 HydrazideTexas RedAlexa Fluor 594590/617A10438
Alexa Fluor 555 HydrazideHydrazide, Tris(triethylammonium)
salt
TRITCAlexa Fluor 555555/5651 mgA20501MP
Alexa Fluor 633 HydrazideTexas RedAlexa Fluor 633632/647A30634
Alexa Fluor 647 HydrazideCy5Alexa Fluor 647650/668A20502
Biocytin
Alexa Fluor 488 Biocytin

Biocytin, sodium salt

FITCAlexa Fluor 488495/519250 mgA12924
Alexa Fluor 594 BiocytinTexas RedAlexa Fluor 594590/617A12922

These conjugates are useful for tissue sections, and cultured cells. They are fixable and photostable retrograde tracers introduced via microinjection or iontophoresis.

Learn more about cholera toxin subunit B

 CarrierStandard filter set(s)FluorophoreEx/Em (nm)QuantityCat. No.
Cholera toxin subunit B
Cholera Toxin Subunit B, Alexa Fluor 488 ConjugateCholera toxin subunit B (Recombinant)FITCAlexa Fluor 488495/519100 µgC34775
Cholera Toxin Subunit B, Alexa Fluor 555 ConjugateTRITCAlexa Fluor 555555/565100 µgC34776
Cholera Toxin Subunit B, Alexa Fluor 594 ConjugateTexas RedAlexa Fluor 594590/617100 µgC34777
Cholera Toxin Subunit B, Alexa Fluor 647 ConjugateCy5Alexa Fluor 647650/668100 µgC34778
Cholera Toxin Subunit B, Biotin-XX ConjugateNABiotin-XXNA100 µgC34779
Cholera Toxin Subunit B, Horseradish Peroxidase ConjugateNAHorseradish peroxidaseNA100 µgC34780
Lipid raft kits
Vybrant Alexa Fluor 488 Lipid Raft Labeling KitCholera toxin subunit B (Recombinant) and anti-cholera toxin subunit B antibody (anti-CT-B)FITCAlexa Fluor 488495/5191 kitV34403
Vybrant Alexa Fluor 555 Lipid Raft Labeling KitTRITCAlexa Fluor 555555/5651 kitV34404
Vybrant Alexa Fluor 594 Lipid Raft Labeling KitTexas RedAlexa Fluor 594590/6171 kitV34405

These cell tracers, also called lipophilic tracers, are useful for tissue sections, in vivo neurons, fixed tissue sections, and cultured cells. They are anterograde and retrograde tracers used for long term studies.

Learn more about DiI and its derivatives

 Standard filter set (s)Ex/Em (nm)FormatFixableQuantityCat. No.
DiI and derivatives
DiI Stain – DiIC18(3)Rhodamine549/565solidNo100 mgD282
DiI Stain – DiIC18(3), crystallinecrystal25 mgD3911
DiD – DiIC18(5)Cy5644/663solid10 mgD7757
DiR – DiIC18(7)Cy5.5750/780solid10 mgD12731
CellTracker DiI
CellTracker CM-DiI Dye, solutionRhodamine549/565solutionYes20 x 50 µgC7000
CellTracker CM-DiI Dyesolid1 mgC7001
Neurotrace DiI
NeuroTrace DiI Tissue-Labeling PasteRhodamine549/565pasteNo500 mgN22880
NeuroTrace CM-DiI Tissue-Labeling PastepasteYes100 mgN22883
Vybrant DiI and derivatives
Vybrant DiO Cell-Labeling SolutionGFP484/501solutionNo1 mLV22886
Vybrant DiI Cell-Labeing SolutionRhodamine549/565solutionNo1 mLV22885
Vybrant CM-DiI Cell-Labeling SolutionsolutionYes1 mLV22888
Vybrant DiD Cell-Labeling SolutionCy5644/663solutionNo1 mLV22887
Vybrant Multicolor Cell-Labeling KitVarioussolutionNo1 mL eachV22889

These cell tracers, also called polar tracers, are useful for tissue sections, in vivo neurons, and cultured cells. They are anterograde and retrograde tracers used to trace neuronal projections and investigate gap junctions.

Learn more about hydrazides and biocytins

 CarrierStandard filter set(s)FluorophoreEx/Em (nm)QuantityCat. No.
Hydrazides
Alexa Fluor 350 HydrazideHydrazide, sodium saltDAPIAlexa Fluor 350346/4425 mgA10439
Alexa Fluor 488 HydrazideFITCAlexa Fluor 488495/519A10436
Alexa Fluor 568 HydrazideRhodamineAlexa Fluor 568578/603A10437
Alexa Fluor 594 HydrazideTexas RedAlexa Fluor 594590/617A10438
Alexa Fluor 555 HydrazideHydrazide, Tris(triethylammonium)
salt
TRITCAlexa Fluor 555555/5651 mgA20501MP
Alexa Fluor 633 HydrazideTexas RedAlexa Fluor 633632/647A30634
Alexa Fluor 647 HydrazideCy5Alexa Fluor 647650/668A20502
Biocytin
Alexa Fluor 488 Biocytin

Biocytin, sodium salt

FITCAlexa Fluor 488495/519250 mgA12924
Alexa Fluor 594 BiocytinTexas RedAlexa Fluor 594590/617A12922

Cholera toxin subunit B

Cholera toxin originates from Vibrio cholerae and is comprised of subunits A and B. Subunit A is an ADP-ribosyltransferase that disrupts protein signaling and will lead to cellular dehydration. Alternatively, subunit B is nontoxic and is involved in the binding of ganglioside GM1 to label lipid rafts (Figure 1). In neurons, cholera toxin subunit B binds to glycosphingolipids in axonal membranes. Introduced via microinjection or iontophoresis, cholera toxin subunit B are fixable and photostable retrograde tracers.

While cholera toxin subunit B Alexa Fluor conjugates offer sensitive and selective binding and neuronal tracing, cholera toxin subunit B biotin-XX and horseradish peroxidase conjugates are available for use with tyramide signal amplification (TSA).

Learn more about cholera toxin b and other protein conjugates

Three images of macrophage stained with red-labeled cholera toxin subunit b or green cholera toxin b antibody, then yellow overlay
Figure 1. J774 mouse macrophage cell stained with Alexa Fluor 555 dye-labeled cholera toxin subunit B. A J774 mouse macrophage cell sequentially stained with BODIPY FL ganglioside GM1 and then with Alexa Fluor 555 dye-labeled cholera toxin subunit B (also available as a component of Vybrant Alexa Fluor 555 Lipid Raft Labeling Kit). The cell was then treated with an anti-CT-B antibody (a component of Vybrant Alexa Fluor 555 Lipid Raft Labeling Kit) to induce crosslinking. Alexa Fluor 555 dye fluorescence (left panel, red) and BODIPY FL dye fluorescence (center panel, green) were imaged separately and overlaid to emphasize the coincident staining (right panel, yellow). Nuclei were stained with blue-fluorescent Hoechst 33258.


DiI and derivatives

DiI and its derivatives are lipophilic tracers, or carbocyanine dyes, that label cell membranes without visibly affecting cell viability. These dyes are weakly fluorescent in water but are highly fluorescent in membranes where they diffuse laterally in the plasma membrane. Lipophilic tracers are useful for live cell staining in tissue sections, in vivo neurons (Figure 2), and cultured cells. Additionally, they can be used in fixed tissue sections, where they can be used to trace neuronal projections (Figure 3). DiI and its derivatives can also be used for long-term studies.

These lipophilic tracers can be loaded via microinjection or iontophoresis and used in both anterograde and retrograde transport studies. They come as crystal and paste preparations which provides convenient options for loading tissues.

Learn more about lipophilic dyes for membrane labeling

Zebrafish brain with green stained nuclei and orange labeled axons in the dorsonasal retina
Figure 2. Tracer DiI as a diagnostic tool to evaluate retinal and tectal patterns in a chimeric five-day-old zebrafish brain. Axons in the dorsonasal retina of a zebrafish embryo labeled with DiI, and nuclei counterstained with SYTOX Green nucleic acid stain. This digital image, previously published on the cover of Development 125 (13) (1999), was created by Alexander Picker and Michael Brand, Neurobiology, University of Heidelberg. The image is used with the permission of The Company of Biologists, Ltd.
Paraformaldehyde-fixed tissue section with green and red labeled neurons from nucleus of thalamus
Figure 3. Confocal microscope image of retrogradely labeled neurons in the ventrobasal nucleus of the thalamus in newborn mouse. Paired dye deposits of DiI (DiI18(3); pseudocolored green) and DiD (DiIC18(5); pseudocolored red) were injected into this 500 µm-thick section, which was then fixed with paraformaldehyde and incubated for two weeks at 37°C to allow retrograde dye transport to proceed. Image contributed by Ariel Agmon, West Virginia University.

Hydrazides and Biocytins

Hydrazides and biocytins are fixable and photostable polar tracers. They have low molecular weights (<1,000 Da) and bright fluorescent signals. Hydrazides and biocytins can be used to investigate cell-cell and cell-liposome fusion. These cell-impermeant polar tracers can also be used to trace neuronal projections and investigate gap junctions (Figures 4, 5). Since they can cross gap junctions, biocytins and hydrazides are often used to investigate neuronal communication along with labeled dextrans, which are retained within the cell (Figure 6). Biocytins can also be detected using fluorescent streptavidin conjugates for signal amplification such as TSA and ELF technologies.

These polar tracers can be loaded using microinjection, pinocytosis, or other techniques that temporarily permeabilize the cell membrane. Additionally, electroporation can be used to transport these tracers into cells.

Learn more about fixable polar tracers

Three images of ganglion cells stained with red or green polar tracers on top and middle, then yellow dual stain on bottom
Figure 4. Rabbit retinal ganglion cells loaded with Alexa Fluor 488 hydrazide and Alexa Fluor 568 hydrazide. Three rabbit retinal ganglion cells injected with red-fluorescent Alexa Fluor 568 hydrazide (top cell), green-fluorescent Alexa Fluor 488 hydrazide (middle cell), or both the Alexa Fluor 488 and the Alexa Fluor 568 hydrazides (bottom cell), where yellow fluorescence indicates the co-localization of the two dyes. Image contributed by Michiel van Wyk, University of Queensland, Australia.
Neurons stained with green and red polar tracers
Figure 5. Neural cells loaded with Alexa Fluor 488 hydrazide and Alexa Fluor 568 hydrazide. Processes of a Leydig neuron iontophoretically filled with green-fluorescent Alexa Fluor 488 hydrazide and the cell body of a giant glial cell filled with red-fluorescent Alexa Fluor 568 hydrazide in a leech ganglion. Image contributed by F. Britz, C. Lohr, J. Schmidt and J.W. Deitmer, University of Kaiserslautern, Germany.
Neurons stained with red polar tracer and green dextran probe shown with co-localization in yellow
Figure 6. Co-stain of neuron with Dextran and Hydrazide. Confocal image stack of a 10,000 MW Calcium Green dextran-labeled climbing fiber in a sagittal cerebellar slice, showing incoming axon and terminal arborization (in yellow). The Purkinje cell innervated by this fiber was labeled with Alexa Fluor 568 hydrazide.

For Research Use Only. Not for use in diagnostic procedures.