Agglutination assay was adapted from Pampana (1933 (link)). Briefly, bacteria were scraped directly from an LB-agar plate into 1 ml of 0.9% NaCl or taken directly from an overnight culture and mixed with 0.5 ml of 0.2% acriflavine (Acriflavine hydrochloride, Sigma A8251). Agglutination was checked after 10–30 min incubation at room temperature. Experiments were performed a minimum of three times and one representative experiment is shown.
Acriflavine hydrochloride
Manufactured by Merck Group
Sourced in Germany
Acriflavine hydrochloride is a synthetic dye that is commonly used as a laboratory reagent. It is a crystalline solid with a yellow to orange color. The compound has a molecular formula of C₁₃H₁₄N₃Cl and a molecular weight of 263.74 g/mol. Acriflavine hydrochloride is soluble in water and is often used in biological and biochemical applications, such as staining and labeling procedures.
Automatically generated - may contain errors
Lab products found in correlation
Streptomycin, by Thermo Fisher Scientific (2 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (2 mentions)
Sybr green 1 solution, by Thermo Fisher Scientific (1 mentions)
Ammonium hydroxide, by BD (1 mentions)
Proflavine hydrochloride, by Merck Group (1 mentions)
Foetal bovine serum (fbs), by Merck Group (1 mentions)
Xvivo system, by Biospherix (1 mentions)
Bcr abl cells, by American Type Culture Collection (1 mentions)
Zinpyr 1, by Cayman Chemical (1 mentions)
Lsm 710 microscope, by Zeiss (1 mentions)
Spc 152, by Becker & Hickl (1 mentions)
Hpm 100 40, by Becker & Hickl (1 mentions)
Potassium hydroxide, by Alpha Chemika (1 mentions)
Sodium hydroxide (naoh), by Alpha Chemika (1 mentions)
6 protocols using acriflavine hydrochloride
1
Bacterial Agglutination Assay
2
Simultaneous Staining of Thraustochytrid Cell Walls and Nuclei
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Acriflavine (3,6-diamino-10-methylacridinium chloride mixture with 3,6-diaminoacridine) was used to concurrently stain thraustochytrid cell walls containing sulfated polysaccharides (red) and the nucleus (yellow-green), as described previously (30 ). Thraustochytrid cultures and seawater subsamples were stained by adding 12 μL acriflavine hydrochloride (Sigma, Germany) solution (5 mg mL−1 in TE buffer) into 3 mL of the sample. After a brief vortex, the resulting solutions were incubated in the dark at room temperature for 30 min. Seawater subsamples for the bacterioplankton analysis were diluted (1:10) with 0.22-μm filtered TE buffer and then stained with 12.5 μL SYBR-I Green solution (1:500 dilution; Molecular Probes, Eugene, OL, USA), followed by an incubation in the dark at room temperature for 10 min (8 (link)). Yellow-green fluorescent polystyrene latex beads with a diameter of 1 μm (Molecular Probes) were added to each FCM sample as an internal standard.
3
Synthesis of Hydroxyapatite and Copper Nanoparticles
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Calcium nitrate tetrahydrate
(Ca(NO3)2·4H2O, Lachema Brno),
ammonium dihydrogen orthophosphate (NH4·H2PO4, ADWIC), ammonium hydroxide (NH4OH, 25%
NH3, BDH AnalaR), and cetyltrimethylammonium bromide (CTAB,
Merck) were used without additional purification in the 1D hydroxyapatite
(1D HAP) synthesis. Copper nitrate trihydrate (Cu(NO3)2·3H2O, assay: 98%, Laboratory Chemicals) and
ascorbic acid (assay: 98%, Alpha Chemicals) were used for the synthesis
of Cu NPs. 4-NP (assay: 98%) and sodium borohydride (NaBH4, assay: 98%) were obtained from Research-Lab Fine Chem Industries
(India) and SDFCL, respectively. Sigma-Aldrich (Germany) provided
the organic dyes (Congo red and acriflavine hydrochloride, ACF).
(Ca(NO3)2·4H2O, Lachema Brno),
ammonium dihydrogen orthophosphate (NH4·H2PO4, ADWIC), ammonium hydroxide (NH4OH, 25%
NH3, BDH AnalaR), and cetyltrimethylammonium bromide (CTAB,
Merck) were used without additional purification in the 1D hydroxyapatite
(1D HAP) synthesis. Copper nitrate trihydrate (Cu(NO3)2·3H2O, assay: 98%, Laboratory Chemicals) and
ascorbic acid (assay: 98%, Alpha Chemicals) were used for the synthesis
of Cu NPs. 4-NP (assay: 98%) and sodium borohydride (NaBH4, assay: 98%) were obtained from Research-Lab Fine Chem Industries
(India) and SDFCL, respectively. Sigma-Aldrich (Germany) provided
the organic dyes (Congo red and acriflavine hydrochloride, ACF).
4
Culturing BCR-ABL+ Leukemia Cell Lines
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K‐562, KU‐812, KCL‐22 and MV‐4‐11 cell lines were obtained from Deutsche Sammlung von Mikroorganismens und Zellkulturen (DSMZ), and IM‐sensitive (K562S) and IM‐resistant (K562R) BCR‐ABL+ cells from American Type Culture Collection (ATCC). Cells were maintained according to the supplier's recommendations. All cell lines were cultured in RPMI 1640 medium containing 2 mmol/L glutamine, 100 U/mL penicillin, 100 µg/mL streptomycin (Gibco, Thermo Fisher, Waltham, MA, USA) and 10% foetal bovine serum (FBS, Thermo Scientific, Sigma‐Aldrich, St Louis, MO, USA) at 37°C, 5% CO2. K562R cells were cultured with 1 µmol/L imatinib mesylate (IM). IM was purchased from Selleckchem (Houston, TX, USA), Acriflavine hydrochloride and proflavine hydrochloride from Sigma‐Aldrich (St‐Louis, MO, USA). Cells were cultured at 1% O2 (Xvivo System, Biospherix, Parish, NY, USA).
5
Multimodal Imaging of Zinc Homeostasis
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Full-thickness skin set aside for en face imaging was stained with acriflavine hydrochloride (Sigma-Aldrich, Castle Hill, Australia) 100 µM in water overnight. Skin cryosections were stained with 10 µM Zinpyr-1 (Cayman Chemical Co., Ann Arbor, MI, USA).
For multiphoton microscopy (MPM), the 488 nm argon laser was used to excite Zinpyr-1, while the 800 nm tuneable titanium-sapphire Mai-Tai was used to excite ZnPT. Emission was collected from the descanned line under a filter (520–560 nm) for single-photon excitation and a non-descanned filter (395–420 nm) for two-photon excitation. A 40X water immersion objective was used.
Fluorescence lifetime imaging microscopy (FLIM) took place along the non-descanned line of the Zeiss LSM710 microscope, fitted with two bh GaAsP hybrid detectors HPM100-40 and two TCSPC modules SPC-152 (Becker & Hickl GmbH, Berlin, Germany). The samples were excited at 740 and 800 nm. Emission was collected using a 405/10 and 540/20 nm bandpass filter (Semrock Inc., Rochester, NY, USA). Images were captured over 5 min. For en face imaging, acriflavine-stained skin was imaged from surface, 40 and 90 µm using the same FLIM parameters as for Zinpyr-1.
Zin-pyr-1 cryosections were first imaged by MPM using the tile-scan function to capture an entire follicle. FLIM images were then acquired along the length of the follicle for detection of ZnPT (λexc = 740 nm).
For multiphoton microscopy (MPM), the 488 nm argon laser was used to excite Zinpyr-1, while the 800 nm tuneable titanium-sapphire Mai-Tai was used to excite ZnPT. Emission was collected from the descanned line under a filter (520–560 nm) for single-photon excitation and a non-descanned filter (395–420 nm) for two-photon excitation. A 40X water immersion objective was used.
Fluorescence lifetime imaging microscopy (FLIM) took place along the non-descanned line of the Zeiss LSM710 microscope, fitted with two bh GaAsP hybrid detectors HPM100-40 and two TCSPC modules SPC-152 (Becker & Hickl GmbH, Berlin, Germany). The samples were excited at 740 and 800 nm. Emission was collected using a 405/10 and 540/20 nm bandpass filter (Semrock Inc., Rochester, NY, USA). Images were captured over 5 min. For en face imaging, acriflavine-stained skin was imaged from surface, 40 and 90 µm using the same FLIM parameters as for Zinpyr-1.
Zin-pyr-1 cryosections were first imaged by MPM using the tile-scan function to capture an entire follicle. FLIM images were then acquired along the length of the follicle for detection of ZnPT (λexc = 740 nm).
6
Synthesis and Characterization of Metal-Dye Nanocomposites
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All materials were of analytical grade and used without any further purification. All solutions were prepared using double-distilled water. Zinc acetate dihydrate (Zn (CH3COO)2·2H2O, assay: 99.5%), sodium hydroxide (NaOH, assay: 99%), potassium hydroxide (KOH, assay: 99%), and l -ascorbic acid (assay: 98%) were purchased from Alpha Chemika. Copper nitrate Cu (NO3)2·3H2O (assay: 98%, Laboratory Chemicals) and 4-nitrophenol (assay: 98%) was supplied by Research-Lab Fine Chem Industries (India), sodium borohydride NaBH4 (assay: 98%) was purchased from SDFCL, while absolute ethanol (ADWIC), methylene blue (MB), Congo red (CR), and acriflavine hydrochloride (ACF) dyes were bought from Sigma-Aldrich (Germany). The chemical structures of 4-NP, MB, CR, and ACF are depicted in Fig. 1 .
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