FluoroGold (也称为Hydroxystilbamidine)，是一款细胞质染料与神经元逆行示踪剂。其核心优势在于能高效进行逆行轴突运输，可实现大量枝晶填充，且荧光亮度高、抗淬灭性强。该产品广泛应用于神经元示踪与组织化学染色，适配压力注射、离子电渗法等多种使用方式，兼容多数常规固定剂，可满足冷冻切片、石蜡切片等不同组织学处理需求，为神经科学研究提供可靠工具。
作为神经科学领域经典的示踪工具，FluoroGold能够精准标记神经投射路径，清晰解析神经元连接网络，为神经环路构建、神经损伤修复机制、神经系统疾病病理研究等方向。凭借优异的抗褪色性和稳定的荧光性能，成为神经元示踪领域不可或缺的核心试剂。
注：效能等同于AAT Bioquest Hydroxystilbamidine(17514)、Biotium Hydroxystilbamidine (Fluoro-Gold™)(80014)；
注：本文中 Fluoro-Gold™为 Fluorochrome, LLC 的商标和注册商标。
应用范围：神经元逆行示踪，组织化学染色，轴突运输，细胞移植与组织工程，病理模型研究等
产品特点：
1、低细胞毒性：对细胞损伤小，保障实验过程中细胞活性与生理状态稳定；
2、高灵敏特异性：低浓度下即可清晰标记胞浆，清晰显示树突分枝细节，且荧光亮度高，无背景干扰；
3、专业逆行示踪：可实现神经通路的逆向投射追踪，适配超短路径至长距离投射的多种实验需求；
4、超长稳定长效：在细胞内分解极慢，动态保证标记强度，完美适配长期追踪实验；
5、多技术多色兼容：荧光染料光谱重叠少，可搭配其他染料进行多色复染，适配多色荧光共标实验。
产品参数：
1、Ex/Em：361/536 nm；
2、CAS 号：223769-64-0；
3、分子式：C18H24N4O7S2；
4、分子量：472.5；
产品组分：
注：以压力注射法，每次使用0.1 μL染料，染色工作液浓度4%计算。
注意事项：
1、对于大多数应用，荧光金染料浓度为2%-4%，在进行无菌实验时，注意对染料进行无菌过滤操作。
2、荧光金几乎可以兼容任何固定剂，也可以不使用固定剂。常用的固定剂为4%多聚甲醛，含有高浓度重金属 (例如：锇，汞)的固定剂会淬灭荧光，而高浓度 (超过1%)的戊二醛可能会增加背景荧光，故最好在实验前进行预实验避免以上因素的影响。
3、含荧光金的组织可以根据任何常见的组织学技术进行处理。这些组织包括未固定组织的冷冻切片 (10 μm)，固定组织的冷冻切片 (20 μm)，或者石蜡切片 (3~10 μm)。
4、对于通过微量注射器或微量移液器进行压力注射，应将荧光金溶于蒸馏水或0.9%的生理盐水中。荧光金也可用作悬浮液于0.2 M中性磷酸盐缓冲液中。但是悬浮的颗粒可能会堵塞微量移液器吸头。对于离子电渗法，在0.1 M醋酸盐缓冲液 (pH 3.3)中配制1%的荧光金溶液。
5、对于注射部位的描述：几乎任何中枢或外周神经系统结构均可注射荧光金来分析逆行转运。在外周神经系统中，可以对神经节和外周目标进行研究。为了研究外周神经，应切断或损伤神经，然后浸入或注射5%的荧光金水溶液。由于完整通道的纤维不会显著吸收荧光金，因此必须切断或严重破坏纤维才能吸收染料。
6、请尽量避光，减缓荧光金的淬灭。
7、使用前请将产品瞬时离心，再进行后续操作。
8、本产品仅限于科研用途并且不得存放于普通住宅内。
9、为了您的安全和健康，请遵循您所在常规实验室安全规定。
使用说明：
一、 实验前准备
试剂准备：将试剂从储存条件 (如4 °C)取出，平衡至室温 (15-25°C)。
二、 操作步骤
荧光金溶于水，现配现用，避免降解。对于大多数实验，荧光金染料浓度为1%-10%即可，初次实验建议浓度为4%，如果在注射部位发生坏死，或者标记过于强烈，可适当降低浓度。
注：不同实验所需荧光金染液浓度需参考文献或通过预实验确定。
1. 使用方法：
(1) 晶体：示踪的晶体可以从微量移液器的尖端注入。
(2) 离子电渗法：4~10 s的脉冲离子疗法 (+5 to+10 μa/10 min)可以产生离散的、小的注射位点。
(3) 压力注射：进样量为0.05~1 μL，通常为0.1~0.2 μL。
2. 固定：(可选)
如需固定选择 4%多聚甲醛的PBS水溶液，建议使用去离子水，溶液中的重金属离子会对染料有淬灭作用，而1%的戊二醛会增加背景。梯度乙醇脱水，再进行脑组织冰冻切片，切20-40 μm厚。
3. 组织化学处理：(可选)
标记荧光金的组织可用于多种组织化学处理，如冰冻组织或固定组织的下游处理。
4. 联合处理办法：(可选)
荧光金组织切片可用第二种其他标记物进行标记，如放射自显影，HRP组织化学、免疫组织化学、以及其他荧光标记物等。
5. 固定：(可选)
实验结束的切片可以用封片剂封片。
6. 检测与成像：
荧光金可直接用荧光显微镜或激光共聚焦进行观察，使用滤光片Ex：361, Em：536 nm。
三、 结果判读
定性分析(显微镜)：
图 1. 皮肤神经元FluoroGold染色图
正常示踪结果：FluoroGold被皮肤神经末梢吸收，沿轴突逆行运输至神经元胞体，标记支配该皮肤区域的神经元。
注：图片引自参考文献《A topical Chinese herbal inhibits pruritus and skin inflammation via neural TRPM8 in atopic dermatitis》（doi: 10.1016/j.phymed.2025.156524.）
常见疑问与解答：
1、问：我用5%浓度的荧光金发现注射部位有坏死，是什么原因？
答：荧光金标记浓度过高导致，可通过设置不同浓度梯度染液注射，确定最佳染料浓度。
2、问：用文献中建议的荧光金染液浓度标记，发现背景荧光很高？
答：排除一下样本是否用了高浓度的戊二醛固定剂，若是，更改为4%多聚甲醛溶液固定测试。
FluoroGold (also known as Hydroxystilbamidine) is a cytoplasmic dye and neuronal retrograde tracer.Its key advantages include efficient retrograde axonal transport, excellent dendritic filling, high fluorescence intensity, and strong photostability.This product is widely used for neuronal tracing and histochemical staining.It is compatible with pressure injection, iontophoresis, and other administration methods, works with most routine fixatives, and supports various histological processing such as frozen sections and paraffin sections, providing a reliable tool for neuroscience research.
As a classic tracer in neuroscience, FluoroGold enables precise labeling of neural projection pathways and clear visualization of neuronal networks.It supports studies on neural circuit assembly, mechanisms of nerve injury and repair, and pathogenesis of neurological diseases.With excellent resistance to photobleaching and stable fluorescence performance, it has become an indispensable core reagent for neuronal tracing.
Note: Performance equivalent to AAT Bioquest Hydroxystilbamidine (17514), Biotium Hydroxystilbamidine (Fluoro-Gold™) (80014).
Note: Fluoro-Gold™ is a trademark and registered trademark of Fluorochrome, LLC.
Applications
Neuronal retrograde tracing, histochemical staining, axonal transport studies, cell transplantation and tissue engineering, pathological model research, etc.
Product Features
1. Low cytotoxicity: Minimal damage to cells, maintaining stable cell viability and physiological status during experiments.
2. High sensitivity and specificity: Clearly labels cytoplasm at low concentrations, reveals fine dendritic branches, with bright fluorescence and low background.
3. Specialized retrograde tracing: Enables retrograde labeling of neural pathways, suitable for both short‑range and long‑range projections.
4. Long‑term stability: Degrades very slowly inside cells, maintains labeling intensity, ideal for long‑term tracing experiments.
5. Multicolor compatibility: Minimal spectral overlap with other fluorophores, supports multiplex fluorescence co‑labeling experiments.
Product Parameters
1. Ex/Em: 361/536 nm
2. CAS No.: 223769-64-0
Molecular Formula: C₁₈H₂₄N₄O₇S₂
Molecular Weight: 472.5
Components
Component
| 1200 T | | FluoroGold | 5 mg |
|
Note: Calculated for pressure injection of 0.1 μL per injection at a working concentration of 4%.
Precautions
1. For most applications, FluoroGold is used at 2%–4%. For sterile experiments, filter the solution aseptically.
2. FluoroGold is compatible with nearly all fixatives and can also be used without fixation. The most common fixative is 4% paraformaldehyde. Fixatives containing high concentrations of heavy metals (e.g., osmium, mercury) quench fluorescence. High concentrations (> 1%) of glutaraldehyde may increase background fluorescence. Pilot testing is recommended before formal experiments.
3. Tissues labeled with FluoroGold can be processed by standard histological techniques: frozen sections of unfixed tissue (10 μm), frozen sections of fixed tissue (20 μm), or paraffin sections (3-10 μm).
4. For pressure injection via syringe or micropipette, dissolve FluoroGold in distilled water or 0.9% saline. It may also be used as a suspension in 0.2 M neutral phosphate buffer, but suspended particles may clog micropipette tips. For iontophoresis, prepare a 1% solution in 0.1 M acetate buffer (pH 3.3).
5. Injection sites: Almost any structure in the central or peripheral nervous system can be injected for retrograde transport analysis. In the peripheral nervous system, ganglia and peripheral targets can be studied. To label peripheral nerves, cut or injure the nerve, then immerse or inject 5% aqueous FluoroGold. Intact fibers do not significantly take up FluoroGold; fibers must be cut or severely damaged.
6. Protect from light to reduce photobleaching.
7. Briefly centrifuge the product before use.
8. For research use only. Do not store in residential areas.
9. Follow standard laboratory safety protocols for your health and safety.
Instructions for Use
I. Pre‑Experiment Preparation
Reagent preparation: Remove the reagent from storage (e.g., 4 °C) and equilibrate to room temperature (15-25 °C).
II. Procedure
FluoroGold is water‑soluble. Prepare fresh solutions to avoid degradation.For most experiments, 1%-10% is suitable; 4% is recommended for initial use.If necrosis at the injection site or excessive labeling occurs, reduce the concentration.
Note: Optimal concentration should be determined from literature or pilot tests.
1. Administration Methods
(1) Crystalline application: Crystals can be applied from micropipette tips.
(2) Iontophoresis: 4–10 s pulses (+5 to +10 μA / 10 min) produce small, discrete injection sites.
(3) Pressure injection: Injection volume 0.05–1 μL, typically 0.1–0.2 μL.
2. Fixation (Optional)
If fixation is needed, use 4% paraformaldehyde in PBS.Deionized water is recommended, as heavy metal ions quench fluorescence; 1% glutaraldehyde increases background.Dehydrate in graded ethanol, then prepare 20–40 μm frozen brain sections.
3. Histochemical Processing (Optional)
Labeled tissues can be processed by standard histology methods, including frozen or fixed tissue protocols.
4. Combined Labeling (Optional)
FluoroGold‑labeled sections can be used with secondary markers: autoradiography, HRP histochemistry, immunohistochemistry, and other fluorescent tracers.
5. Coverslipping (Optional)
Sections may be mounted with mounting medium after experimentation.
6. Detection & Imaging
Observe directly with fluorescence microscopy or confocal laser microscopy using filters Ex: 361 nm, Em: 536 nm.
III. Interpretation of Results
Qualitative analysis (microscopy):
Figure 1. FluoroGold staining of cutaneous neuronsNormal tracing result: FluoroGold is taken up by cutaneous nerve terminals, transported retrogradely along axons to neuronal somata, labeling neurons innervating the skin region.
Note: Image adapted from reference: A topical Chinese herbal inhibits pruritus and skin inflammation via neural TRPM8 in atopic dermatitis (doi: 10.1016/j.phymed.2025.156524.).
FAQ
1. Q: I see necrosis at the injection site using 5% FluoroGold. Why?
A: Caused by excessively high tracer concentration.Test a concentration gradient to determine the optimal working concentration.
2. Q: Background fluorescence is very high even at the recommended concentration. Why?
A: Check if high‑concentration glutaraldehyde was used for fixation.If so, switch to 4% paraformaldehyde and retest.
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