Hydroxystilbamidine CAS 223769-64-0

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Physical properties

Molecular weight	472.54
Solvent	Water

Spectral properties

Excitation (nm)	358
Emission (nm)	433

Storage, safety and handling

Certificate of Origin	Download PDF
H-phrase	H303, H313, H340
Hazard symbol	T
Intended use	Research Use Only (RUO)
R-phrase	R20, R21, R68
Storage	Freeze (< -15 °C); Minimize light exposure
UNSPSC	41116134

Overview SDS Protocol

CAS

223769-64-0

Molecular weight

472.54

Excitation (nm)

358

Emission (nm)

433

Hydroxystilbamidine (also called Fluoro Gold) is used for staining DNA and RNA. It exhibits distinctively different fluorescence emission profiles when bound to DNA compared to RNA. This cationic dye is also frequently used as a retrograde neuronal tracer. Fluoro-Gold is a trademark of Fluorochrome, Inc.

Example protocol

AT A GLANCE

Biological Applications

Spectral Properties

Excitation = 360 nm; Emission = 625 nm

PREPARATION OF WORKING SOLUTION

Hydroxystilbamidine working solution

1 to 10% Hydroxystilbamidine in water has been successfully used. Initially, a 4% concentration is advised. If undesirable necrosis occurs at the injection site, or labeling is too intense, reduce the concentration to a 2% solution.

SAMPLE EXPERIMENTAL PROTOCOL

The use of Hydroxystilbamidine is essentially the same as other fluorescent tracers. The main difference is that Fluoro-Gold is more flexible in terms of post-injection survival times, concentration range, tissue treatment and compatibility with other histochemical techniques. It is also more resistant to fading, brighter and more permanent than most other fluorescent tracers.

Dye Administration

Pressure injection - This is probably the most frequently used mode of application. Volumes injected range from 0.05-1 µL, typically 0.1-0.2 µL.
Crystal - A crystal of the tracer can be administered from the tip of a micro-pipette.

Fixation

Although any fixative, or no fixative, can be used, PBS containing 4% formaldehyde is most frequently employed. Fixatives containing high concentrations of heavy metals (eg. osmium, mercury) will quench the fluorescence, while high concentrations (over 1%) of glutaraldehyde may increase background fluorescence

Histochemical Processing

Tissue containing Hydroxystilbamidine may be processed according to virtually any common histological technique. Frozen sections of fixed tissue are most frequently used.

Combined Methods

At this point of processing, sections may be further processed for a second marker such as autoradiography, HRP histochemistry, immunocytochemistry, a second fluorescent tracer, fluorescent counterstain, etc.

Mounting, Clearing and Coverslipping

Sections are typically mounted on gelatin-coated slides, air-dried, immersed in xylene, and coverslipped with nonfluorescent DPX plastic mounting media. Sections may be dehydrated with graded alcohols, unless this is not compatible with a second tracer. If Hydroxystilbamidine is to be combined with fluorescence immunocytochemistsry, then sections are air-dried and directly coverslipped with DPX.

Examination and Photography

Hydroxystilbamidine can be visualized with a fluorescence microscope using a wide band ultraviolet excitation filter (excitation - 323 nm, emission - 620 nm at neutral pH). A gold color is emitted when tissue has been processed with neutral pH buffer, whereas a blue color is emitted when tissue is processed with acidic (eg. PH 3.3) pH buffer. It can be photographed digitally or with film (use Ektachrome 200-400 ASA film for color prints and comparable speed film for black and white prints). Most exposure times range from 10-60 second exposures, depending on the objective magnification and the intensity of the label. Thirty (30) second exposures are about average. Multiple exposures may be exploited to simultaneously visualize Hydroxystilbamidine and another tracer. Thus, UV would be combined with bright field illumination to simultaneously locate Fluoro-Gold with HRP or silver grains in autoradiography. Similarly, blue light excitation can be combined to also visualize the green emission color of FITC, while green excitation light may be used to simultaneously observe the red emission color of propidium iodide, or ethidium bromide (a fluorescent counterstain).

Calculators

Common stock solution preparation

Table 1. Volume of Water needed to reconstitute specific mass of Hydroxystilbamidine *CAS 223769-64-0* to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.

	0.1 mg	0.5 mg	1 mg	5 mg	10 mg
1 mM	211.622 µL	1.058 mL	2.116 mL	10.581 mL	21.162 mL
5 mM	42.324 µL	211.622 µL	423.245 µL	2.116 mL	4.232 mL
10 mM	21.162 µL	105.811 µL	211.622 µL	1.058 mL	2.116 mL

Molarity calculator

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Spectrum

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Spectral properties

Excitation (nm)	358
Emission (nm)	433

Images

Figure 1. Chemical structure for Hydroxystilbamidine *CAS 223769-64-0*

Citations

View all 4 citations: Citation Explorer

A missense mutation in the MLKL brace region promotes lethal neonatal inflammation and hematopoietic dysfunction
Authors: Hildebrand, Joanne M and Kauppi, Maria and Majewski, Ian J and Liu, Zikou and Cox, Allison J and Miyake, Sanae and Petrie, Emma J and Silk, Michael A and Li, Zhixiu and Tanzer, Maria C and others,
Journal: Nature communications (2020): 1--16

Missense mutations in the MLKL ‘brace’region lead to lethal neonatal inflammation in mice and are present in high frequency in humans
Authors: Hildebrand, Joanne M and Kauppi, Maria and Majewski, Ian J and Liu, Zikou and Cox, Allison and Miyake, Sanae and Petrie, Emma J and Silk, Michael A and Li, Zhixiu and Tanzer, Maria C and others,
Journal: bioRxiv (2019): 628370

Membrane-Associated RING-CH (MARCH) proteins down-regulate cell surface expression of the interleukin-6 receptor alpha chain (IL6Ra)
Authors: Babon, Jeffrey J and Stockwell, Dina and DiRago, Ladina and Zhang, Jian-Guo and Laktyushin, Artem and Villadangos, Jose and Ching, Alan and Ishido, Satoshi and Hilton, Douglas J and Alex, undefined and er, Warren S and others, undefined
Journal: Biochemical Journal (2019): BCJ20190577

Inoculation of α-synuclein preformed fibrils into the mouse gastrointestinal tract induces Lewy body-like aggregates in the brainstem via the vagus nerve
Authors: Uemura, Norihito and Yagi, Hisashi and Uemura, Maiko T and Hatanaka, Yusuke and Yamakado, Hodaka and Takahashi, Ryosuke
Journal: Molecular Neurodegeneration (2018): 21

References

View all 22 references: Citation Explorer

Retrogradely transported fluorogold accumulates in lysosomes of neurons and is detectable ultrastructurally using post-embedding immuno-gold methods
Authors: Persson S, Havton LA.
Journal: J Neurosci Methods (2009): 42

Fluorogold induces persistent neurological deficits and circling behavior in mice over-expressing human mutant tau
Authors: He Z., undefined
Journal: Curr Neurovasc Res (2009): 54

A morphologic study of Fluorogold labeled tensor tympani motoneurons in mice
Authors: Mukerji S, Brown MC, Lee DJ.
Journal: Brain Res (2009): 59

Organisation of the catecholaminergic system in the vagal motor nuclei of pigs: a retrograde fluorogold tract tracing study combined with immunohistochemistry of catecholaminergic synthesizing enzymes
Authors: Chaillou E, Tillet Y, Malbert CH.
Journal: J Chem Neuroanat (2009): 257

Fluorogold labeling of descending brain neurons in larval lamprey does not cause cell death
Authors: McClellan AD, Zhang L, Palmer R.
Journal: Neurosci Lett (2006): 119

Improved detection of fluorogold-labeled neurons in long-term studies
Authors: Akhavan M, Hoang TX, Havton LA.
Journal: J Neurosci Methods (2006): 156

Origins of lateral hypothalamic afferents associated with N-methyl-d-aspartic acid-elicited eating studied using reverse microdialysis of NMDA and Fluorogold
Authors: Duva MA, Tomkins EM, Mor and a LM, Kaplan R, Sukhaseum A, Stanley BG.
Journal: Neurosci Res (2005): 95

Tracer coupling of neurons in the rat retina inner nuclear layer labeled by Fluorogold
Authors: Abdel-Majid RM, Archibald ML, Tremblay F, Baldridge WH.
Journal: Brain Res (2005): 114

Deficient motor innervation of the sphincter mechanism in fetal rats with anorectal malformation: a quantitative study by fluorogold retrograde tracing
Authors: Yuan ZW, Lui VC, Tam PK.
Journal: J Pediatr Surg (2003): 1383

Anatomy of olivocochlear neurons in the hamster studied with FluoroGold
Authors: Sanchez-Gonzalez MA, Warr WB, Lopez DE.
Journal: Hear Res (2003): 65