Nile Blue A (sulfate)

Names

[ Name ]:
Nile Blue A (sulfate)

[Synonym ]:
Bis[N-(5-amino-9H-benzo[a]phenoxazin-9-ylidene)-N-ethylethanaminium] sulfate
EINECS 222-832-5
Nile Blue A sulfate
Bis[9-(diethylamino)-5H-benzo[a]phenoxazin-5-iminium] sulfate
MFCD00064529
Nile Blue A
Basic Blue 12,Nile blue sulfate
Nile Blue A,certified
Nile blue, sulfate
Basic Blue-12
Basic Blue 12 Nile blue sulfate
Nile blue sulfate
Nile Blue A (sulfate)

Biological Activity

[Description]:

Nile Blue A is used to differentiate melanins and lipofuscins. It is also useful for staining fats and preparation of an amperometric glucose sensor.

[Related Catalog]:

Signaling Pathways >> Others >> Others

Dye Reagents

Research Areas >> Others

[In Vitro]

Nile blue A is a basic oxazine dye which is soluble in water and ethyl alcohol. Nile blue A is a satisfactory stain for PHB granules in bacteria and is in fact superior to Sudan black B for this purpose. Poly-p3-hydroxybutyrate granules exhibits a strong orange fluorescence when stained with Nile blue A. Nile blue A appears to stain many more PHB granules than Sudan black B does and is not as easily ished from the cell by decolorization procedures[1]. Nile blue A is used as a stain for polyhydroxyalkanoic acid-accumulating microorganisms or to detect polyhydroxyalkanoic acids in microorganisms. Escherichia coli cells that do not accumulate detectable polyhydroxyalkanoic acids can be stained with Nile blue A and that this staining is sufficient for identifying these cells in fluorescence-activated cell sorting (FACS) experiments. Nile blue A staining does not affect either surface display of peptides or specific labeling of these peptides by a second fluorescence. Staining E. coli for flow cytometry using Nile blue A is an easy-to-handle and low-cost alternative to other fluorescent dyes or the intracellular expression of, for example, green fluorescent protein[2]. Nile blue A is one of the most studied benzophenoxazine dyes, as a potent photosensitizer for photodynamic therapy. The dye when administered intravenously disperses throughout the body by circulating through blood and is taken up by most cells that emphasize its interaction with various biomolecule[3].

[Cell Assay]

1% aqueous solution of Nile blue A is prepared and filtered before use. Mild heating may be necessary to fully dissolve the stain. Heat-fixed smears of bacterial cells are stained with the Nile blue A solution at 55°C for 10 min in a coplin staining jar. After being stained, the slides are washed with tap water to remove excess stain and with 8% aqueous acetic acid for 1 min. The stained smear is washed and blotted dry with bibulous paper, remoistened with tap water, and covered with a no. 1 glass cover slip. The preparation is examined with a Nikon Labphot microscope with an episcopic fluorescence attachment[1]. The PHA− strain Escherichia coli UT5600(DE3) is stained with Nile blue A. In an Erlenmeyer flask, 20 mL of Luria–Bertani (LB) broth is inoculated with one colony of UT5600(DE3) and incubated for 14 h at 37 °C and 200 rpm. Subsequently, 20 mL of LB broth containing Nile blue A in a final concentration of 0.5 μg/mL is inoculated with 200 μL of the 14-h culture and cultured to an optical density at 578 nm (OD578) of 0.6. As a control, 20 mL of LB broth (without Nile blue A) is inoculated with 200 μL of the 14-h culture and is also cultured to an optical density at OD578 of 0.6. Every 20 min, the OD578 is determined for both cultures to verify whether there is any influence of the dye on the growth of the bacteria[2]. Nile blue A stock solution is prepared using ethanol as the solvent. The concentration of NB is maintained at 5 μM for all the studies. The solutions are left for 1 h to achieve equilibrium before spectral measurements. The absorption spectra are recorded using Shimadzu Spectrophotometer (UV-1800) and the emission spectra are recorded using Jobin–Yvon Spectrofluorimeter. A 450 nm nano-LED is used as the light source and the fluorescence lifetime is collected at λem=672 nm[3].

[References]

[1]. Ostle AG, et al. Nile blue A as a fluorescent stain for poly-beta-hydroxybutyrate. Appl Environ Microbiol. 1982 Jul;44(1):238-41.

[2]. Betscheider D, et al. Nile blue A for staining Escherichia coli in flow cytometer experiments. Anal Biochem. 2009 Jan 1;384(1):194-6.

[3]. Mishra SS, et al. Spectroscopic investigation of interaction of Nile Blue A, a potent photosensitizer, with bile salts in aqueous medium. J Photochem Photobiol B. 2014 Dec;141:67-75.

[Related Small Molecules]

Chemical & Physical Properties

[ Boiling Point ]:
487.9ºC

[ Melting Point ]:
>300ºC (dec.)(lit.)

[ Molecular Formula ]:
C₂₀H₂₀N₃O₃S_0.5

[ Molecular Weight ]:
366.42

[ Flash Point ]:
248.8ºC

[ PSA ]:
198.76000

[ LogP ]:
7.86520

[ Water Solubility ]:
Soluble in water, ethanol

MSDS

Click to get detail MSDS

Toxicological Information

CHEMICAL IDENTIFICATION

RTECS NUMBER :: DJ1931000

CAS REGISTRY NUMBER :: 3625-57-8

LAST UPDATED :: 199512

DATA ITEMS CITED :: 3

MOLECULAR FORMULA :: C40-H40-N6-O2.O4-S

MOLECULAR WEIGHT :: 732.92

WISWESSER LINE NOTATION :: T D6 B666 CN JOJ GN2&2 MZ 2 &.S-O4 &15/33

HEALTH HAZARD DATA

ACUTE TOXICITY DATA

TYPE OF TEST :: LD - Lethal dose

ROUTE OF EXPOSURE :: Intraperitoneal

SPECIES OBSERVED :: Rodent - mouse

DOSE/DURATION :: >500 mg/kg

TOXIC EFFECTS :: Details of toxic effects not reported other than lethal dose value

REFERENCE :: CBCCT* "Summary Tables of Biological Tests," National Research Council Chemical-Biological Coordination Center. (National Academy of Science Library, 2101 Constitution Ave., NW, Washington, DC 20418) Volume(issue)/page/year: 4,46,1952

TYPE OF TEST :: LDLo - Lowest published lethal dose

ROUTE OF EXPOSURE :: Intravenous

SPECIES OBSERVED :: Rodent - mouse

DOSE/DURATION :: 27 mg/kg

TOXIC EFFECTS :: Details of toxic effects not reported other than lethal dose value

REFERENCE :: TXAPA9 Toxicology and Applied Pharmacology. (Academic Press, Inc., 1 E. First St., Duluth, MN 55802) V.1- 1959- Volume(issue)/page/year: 44,225,1978 *** NIOSH STANDARDS DEVELOPMENT AND SURVEILLANCE DATA *** NIOSH OCCUPATIONAL EXPOSURE SURVEY DATA : NOES - National Occupational Exposure Survey (1983) NOES Hazard Code - X4351 No. of Facilities: 6 (estimated) No. of Industries: 1 No. of Occupations: 1 No. of Employees: 39 (estimated) No. of Female Employees: 20 (estimated)

Safety Information

[ Personal Protective Equipment ]:
Eyeshields;Gloves;type N95 (US);type P1 (EN143) respirator filter

[ Hazard Codes ]:
Xi: Irritant;

[ Safety Phrases ]:
S24/25

[ RIDADR ]:
NONH for all modes of transport

[ WGK Germany ]:
3

[ RTECS ]:
DJ1931000

[ HS Code ]:
2934999090

Customs

[ HS Code ]: 2934999090

[ Summary ]:
2934999090. other heterocyclic compounds. VAT:17.0%. Tax rebate rate:13.0%. . MFN tariff:6.5%. General tariff:20.0%

Articles

Heterogeneous immunoassay for soy protein determination using nile blue-doped silica nanoparticles as labels and front-surface long-wavelength fluorimetry.

Anal. Chim. Acta 701(2) , 194-9, (2011)

A long-wavelength fluoroimmunoassay for the determination of soy protein is reported for the first time using a conjugate composed of anti-soy protein antibodies bound to nile blue-doped silica nanopa...

Stepwise fluorescence changes of quantum dots: single-molecule spectroscopic studies on the properties of turn-on quantum dots.

Chem. Commun. (Camb.) 48(5) , 723-5, (2012)

Single-molecule spectroscopy of turn-on quantum dots induced by NADPH-dependent biocatalyzed transformations reveals that the fluorescence intensities of quantum dots functionalized with Nile Blue are...

Rapid separations of nile blue stained microorganisms as cationic charged species by chip-CE with LIF.

Electrophoresis 33(9-10) , 1421-6, (2012)

Rapid detection of microorganisms by alternative methods is desirable. Electromigration separation methods have the capability to separate microorganisms according to their charge and size and laser-i...