Carbol fuchsin is a histological stain used in microbiology to differentiate acid-fast bacteria from non-acid-fast bacteria. It is a mixture of basic fuchsin, phenol, and water and is widely used in the Ziehl-Neelsen staining technique for the detection of tuberculosis and other mycobacterial infections. Carbol fuchsin stains the cell walls of acid-fast bacteria bright red, while other cells are unstained or slightly stained. This makes it easier to observe and identify these microorganisms under a microscope. Carbol fuchsin is also used in veterinary medicine and phytopathology for similar purposes.

Physicochemical Properties

Molecular Formula	C21H22CLN3
Molecular Weight	351.87
Exact Mass	431.176
CAS #	4197-24-4
PubChem CID	5463951
Appearance	Typically exists as solid at room temperature
Density	0.99 g/mL at 20ºC
Boiling Point	573.5ºC at 760 mmHg
Flash Point	300.6ºC
LogP	7.563
Hydrogen Bond Donor Count	4
Hydrogen Bond Acceptor Count	3
Rotatable Bond Count	2
Heavy Atom Count	25
Complexity	576
Defined Atom Stereocenter Count	0
SMILES	CC1=CC(=CC=C1N)C(=C2C=CC(=N)C=C2)C3=CC=C(C(=C3)C)N.Cl
InChi Key	HZLHRDBTVSZCBS-GHTYLULLSA-N
InChi Code	InChI=1S/C21H21N3.ClH/c1-13-11-16(5-9-19(13)23)21(15-3-7-18(22)8-4-15)17-6-10-20(24)14(2)12-17;/h3-12,23H,22,24H2,1-2H3;1H/b21-16-,23-19?;
Chemical Name	4-[(Z)-(4-aminophenyl)-(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl]-2-methylaniline;hydrochloride
Synonyms	Carbol Fuchsin; Carbol-Fuchsin; 4197-24-4; Carbol-Fuchsin [USAN]; 8052-17-3; Carbolfuchsin; 4-((4-Aminophenyl)(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl)-2-methylaniline hydrochloride; 4-[(E)-(4-aminophenyl)-(4-imino-3-methylcyclohexa-2,5-dien-1-ylidene)methyl]-2-methylaniline;hydrochloride;
HS Tariff Code	2934.99.9001
Storage	Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month
Shipping Condition	Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Biological Activity

Targets	Biochemical assay reagent; histological stain
ln Vitro	A biochemical reagent called carbolic fuchsin can be utilized in life science research as an organic substance or biomaterial.
ln Vivo	Injection of high-viscosity fluids into subcutaneous tissues may lead to a granulomatous reaction called sclerosing lipogranuloma (SL). Poly-(d,l-lactide-co-glycolide) (PLG or PLGA) microspheres are used as vehicles for extended-release drugs. Here we describe the histopathologic features of a case of SL induced by exenatide extended-release injections, and the staining pattern of PLG microspheres and microsphere remnants with carbol fuchsin [1].
Enzyme Assay	Carbol fuchsin is an acid-fast, lipophilic, magenta-colored dye, often used to identify waxy mycobacterial cell walls.5 Specifically, carbol fuchsin is prepared from phenol and basic fuchsin. Because basic fuchsin is highly soluble in lipids such as mycolic acid, carbol fuchsin is used as a component in the Ziehl-Neelsen and Fite stains in identifying acid-fast bacterial species. Individual PLG spherules combine to create an amalgam in vivo, which is then nonenzymatically hydrolyzed in a process that will continue regardless of the microenvironment, although degradation of the polymers will itself generate an acidic environment within the amalgam.7 Fused, semi-degraded spherules appearing as vacuolated material probably account for the large area of fuchsin-staining in this case. For comparison, Fite stain was also applied to three cases of fat necrosis, including traumatic fat necrosis after excision, lipomembranous fat necrosis, and encapsulated fat necrosis. Staining with carbol fuchsin was not identified in tissue sections of these patterns of fat necrosis [1].
References	[1]. Carbol fuchsin stain enhances detection of poly-(d, l-lactide-co-glycolide) microspheres in exenatide extended-release cutaneous injection-site foreign body reaction. J Cutan Pathol. 2021 Dec;48(12):1520-1522.
Additional Infomation	Coloring Agents: Chemicals and substances that impart color including soluble dyes and insoluble pigments. They are used in INKS; PAINTS; and as INDICATORS AND REAGENTS. A 57-year-old woman with type 2 diabetes mellitus presented for evaluation of painless subcutaneous abdominal masses at the injection sites of exenatide extended-release microspheres. There were two ill-defined, soft and mobile nodules on clinical examination, and the differential included lipomas and angiolipomas. Considering the benign nature of these lesions, it was decided to forego intervention in favor of self-monitoring. On examination at a follow-up appointment 15 months later, there were five non-tender, soft, mobile subcutaneous abdominal nodules. Two of the nodules were excised and the tissue was sent for histopathologic examination to determine the nature of the reaction to the medication injection and to rule out infection. Both specimens showed cystic spaces in the dermis and subcutaneous fat that were surrounded by palisading macrophages and multinucleated giant cells, which were then surrounded by lymphocytes and interstitial fibrosis (Figure 1). Scattered, single, vacuolated giant cells were present, as well as groups of giant cells surrounding large vacuoles (Figure 2). PAS, Gram stain, and acid-fast stains were negative for fungi, bacteria, and acid-fast bacilli. The histopathologic findings were consistent with sclerosing lipogranuloma. Notably, there was a slightly basophilic, vacuolated material present in the cavities and vacuoles (Figure 3), which positively stained with carbol fuchsin (as part of the Fite acid-fast stain) but not with Alcian blue/periodic acid-Schiff (PAS). (Figures 4 and 5) Tiny spheres were visible in the carbol fuchsin-positive material, representing PLG microsphere remnants (Figure 6). Occasional scattered polarizable material was present, which was consistent with residual, intact microspheres.[1]

Solubility Data

Solubility (In Vitro)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Solubility (In Vivo)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples. Injection Formulations (e.g. IP/IV/IM/SC) Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] *Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline) Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline) Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline) Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil) Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Oral Formulations Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). Oral Formulation 3: Dissolved in PEG400 Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose Oral Formulation 6: Mixing with food powders Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.8420 mL	14.2098 mL	28.4196 mL
	5 mM	0.5684 mL	2.8420 mL	5.6839 mL
	10 mM	0.2842 mL	1.4210 mL	2.8420 mL

*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.