Enhancing Fluorogold-based neural tract tracing.
Fluorogold (FG) is used by many groups to retrogradely trace nervous system pathways. Fluorogold, while a robust tracer, also is neurotoxic and causes tissue damage at the injection site and leads to motor deficits.



In the current study, we describe a method for enhancing FG-uptake using Triton™ and an overall procedure for reducing FG-related tissue damage while still allowing effective quantification.



Triton™ decreases the amount of FG, as well as the time required for long-distance transport from the thoracic spinal cord to the motor cortex by >4 fold when this distance is >10in. Although small FG concentrations and injection volumes are ideal for minimizing associated tissue damage and motor deficits, they result in difficult-to-detect fluorescence. This can be solved using FG antiserum paired with an ABC chromogen reaction. This ABC chromogen reaction product can remain stable for at least 9 years.



This study is the first to collectively address FG-induced tissue damage and describe methods for minimizing this damage.



Triton™ enhances the uptake of FG in the nervous system, reduces the FG required, and allows for a substantial decrease in tracing time that limits FG-induced motor deficits. Small FG concentration and volume decreases tissue damage but also decreases FG fluorescent detection. Detection challenges are resolved using FG anti-serum and chromogen reactions.
Department of Rehabilitation Medicine, University of Washington, Seattle, WA 98195, United States.
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Mondello
SensoMotoric Instruments, Inc., Boston, MA 02110, United States.
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Jefferson
Kentucky Spinal Cord Injury Research Center, University of Louisville, Louisville, KY, United States; Department of Neurological Surgery, University of Louisville, Louisville, KY 40292, United States; Robley Rex VA Medical Center, Louisville, KY 40206, United States.
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O'Steen
Kentucky Spinal Cord Injury Research Center, University of Louisville, Louisville, KY, United States; Department of Neurological Surgery, University of Louisville, Louisville, KY 40292, United States; Department of Anatomical Sciences and Neurobiology, University of Louisville, Louisville, KY 40292, United States; Department of Bioengineering, University of Louisville, Louisville, KY 40292, United States; Robley Rex VA Medical Center, Louisville, KY 40206, United States. Electronic address: drhowl01@louisville.edu.
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Howland
D000818:Animals; D048588:Benzoxazines; D002415:Cats; D005260:Female; D007150:Immunohistochemistry; D008856:Microscopy, Fluorescence; D009044:Motor Cortex / Q000473:pathology; D009336:Necrosis / Q000209:etiology / Q000473:pathology; D018541:Neptune; D056347:Neuroanatomical Tract-Tracing Techniques; D056325:Neuronal Tract-Tracers / Q000009:adverse effects; D013116:Spinal Cord / Q000166:cytology / Q000000981:diagnostic imaging / Q000473:pathology; D013119:Spinal Cord Injuries / Q000473:pathology; D013266:Stilbamidines / Q000009:adverse effects; D013997:Time Factors
C049774:2-hydroxy-4,4'-diamidinostilbene, methanesulfonate salt; D048588:Benzoxazines; D056325:Neuronal Tract-Tracers; D013266:Stilbamidines; C028911:cresyl violet
Detergent; Fluorogold; Immunohistochemistry; Neurotoxic; Retrograde tracing; Triton