AsPC-1-GFP (1×106 cells) cells were cultured for 3 days. Dishes were washed with PBS and incubated with RPMI-1640 medium with 1 mM 5-ALA hydrochloride (Wako Pure Chemical Industries) for 3 h. Then, we observed the fluorescence of PpIX (excitation, 440 nm; emission, 575–675 nm) and GFP (excitation, 488 nm; emission, 500–560 nm) with an inverted microscope (IX81) equipped with a confocal scanning system (FV1000; both Olympus, Tokyo, Japan).
5 ala hydrochloride
Manufactured by Fujifilm
Sourced in Japan
5-ALA hydrochloride is a chemical compound used in various laboratory applications. It is the hydrochloride salt of 5-aminolevulinic acid, a precursor in the biosynthesis of heme and other tetrapyrrole compounds. The product is commonly used in research and development settings, but a detailed description of its core function is not available while maintaining an unbiased and factual approach.
Automatically generated - may contain errors
Lab products found in correlation
Fv1000, by Olympus (1 mentions)
Fetal bovine serum (fbs), by Fujifilm (1 mentions)
Propidium iodide, by Fujifilm (1 mentions)
Sodium hydroxide (naoh), by Fujifilm (1 mentions)
Phosphate buffered saline (pbs), by Fujifilm (1 mentions)
N n dimethylformamide (dmf), by Fujifilm (1 mentions)
Penicillin, by Fujifilm (1 mentions)
Streptomycin, by Fujifilm (1 mentions)
Isopropanol, by Fujifilm (1 mentions)
Rpmi 1680 medium, by Fujifilm (1 mentions)
Dna damage detection kit containing γh2ax monoclonal antibody and secondary antibody, by Dojindo Laboratories (1 mentions)
Aminophenyl fluorescein apf, by Goryo Chemical (1 mentions)
D luciferin, by Fujifilm (1 mentions)
Synergy h1, by Agilent Technologies (1 mentions)
Szx12, by Olympus (1 mentions)
Hq430lp, by Chroma Technology (1 mentions)
Gfpa cube, by Olympus (1 mentions)
U lh100hg, by Olympus (1 mentions)
6 protocols using 5 ala hydrochloride
1
PpIX and GFP Fluorescence Imaging
2
Mitochondrial Dysfunction and DNA Damage Assays
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5-ALA hydrochloride, RPMI 1680 medium, penicillin, streptomycin, fetal bovine serum (FBS), phosphate-buffered saline (PBS), NaOH, N,N-dimethylformamide, isopropanol, propidium iodide (PI), RNase, and D-luciferin were purchased from Wako Chemicals (Osaka, Japan). A modified Lowry protein assay kit was purchased from Pierce (Rockford, IL, USA). Aminophenyl fluorescein (APF) was purchased from Goryo Chemical (Sapporo, Japan). A DNA damage detection kit (containing γH2AX monoclonal antibody and secondary antibody) was purchased from Dojindo Laboratories (Kumamoto, Japan). A MitoPT® TMRE Mitochondrial Depolarization Assay Kit was purchased from ImmunoChemistry Technology (Bloomington, MN, USA).
3
5-ALA uptake measurement in cells
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RGK-WT and RGK-XRR were seeded at 2 × 104 cells/well in 24-well cell culture plates, respectively, and cultured for overnight. The medium was replaced with 1 mM 5-ALA hydrochloride (FUJIFILM Wako Pure Chemical Corporation) and incubated in the dark for 6 h. After removing the medium and washing three times with PBS, the cells were lysed in 100 μl/well of RIPA buffer and transferred to a 96-well cell culture plate. The fluorescence intensity of the cell lysate was measured with a Synergy H1 microplate reader (BioTek Instruments Inc., Winooski, VT). The wavelength was excited at 415 nm and the fluorescence wavelength at 625 nm was measured.
4
5-ALA Effects on Liver Function in Mice
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Recently, 5-ALA has received much attention as a therapeutic agent [8 , 11 (link)] and feed additive [10 (link)]. Metabolomic analysis revealed that 5-ALA levels
were significantly higher in the portal and abdominal veins of LAO1 KO mice. Consequently,
we focused our efforts on understanding 5-ALA and its effects on the liver. Two
experiments were conducted using WT animals to understand the effects of this chemical on
liver function. For the low-dose experiment, 10 mice were injected daily with saline or 40
mg/kg BW 5-ALA hydrochloride (Fujifilm Wako Pure Chemical Corp., Osaka, Japan) for one
week. The other high-dose experiment involved IP injection of a higher dose of 5-ALA (100
mg/kg BW) or saline every three days for ten days. The doses used in this study were
selected based on the therapeutic doses previously used in mice. After sacrificing the
mice under general anesthesia, the samples were collected.
were significantly higher in the portal and abdominal veins of LAO1 KO mice. Consequently,
we focused our efforts on understanding 5-ALA and its effects on the liver. Two
experiments were conducted using WT animals to understand the effects of this chemical on
liver function. For the low-dose experiment, 10 mice were injected daily with saline or 40
mg/kg BW 5-ALA hydrochloride (Fujifilm Wako Pure Chemical Corp., Osaka, Japan) for one
week. The other high-dose experiment involved IP injection of a higher dose of 5-ALA (100
mg/kg BW) or saline every three days for ten days. The doses used in this study were
selected based on the therapeutic doses previously used in mice. After sacrificing the
mice under general anesthesia, the samples were collected.
5
In Vivo Fluorescence Imaging of Metastatic Tumors
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An aliquot of 1×106 EGFP tagged HT-29 cells was injected into the peritoneal cavity of mice under general anesthesia. After 2 weeks, the mice were intraperitoneally injected with 5-ALA hydrochloride (Wako Pure Chemical Industries, Osaka, Japan) at a dose of 250 mg/kg body weight. Six hours after 5-ALA administration, the mice were euthanized and laparotomy was performed. Metastatic nodules in the omentum were observed in white light and fluorescence images. Fluorescence observation was performed with a stereoscopic microscope (SZX12; Olympus, Tokyo, Japan) equipped with a color CCD digital camera (DP71, Olympus) and a mercury lamp (U-LH100HG; Olympus). We used a spectral analytic system composed of a stereoscopic microscope equipped with an intensified multi-channel spectrophotometer (MCPD-7000, Otsuka Electronics, Osaka, Japan) for spectral analysis. PpIX images (>430 nm, HQ430LP, Chroma Technology Corp., Rockingham, VT, USA) were acquired by exciting at 405±20 nm (D405/20x, Chroma Technology Corp.), and EGFP fluorescence images (510–530 nm) (GFPA cube, Olympus) were acquired by exciting at 460–490 nm (GFPA cube, Olympus); all images were recorded in the red, green and blue format.
6
Fluorescence Imaging of 5-ALA Tumor Uptake
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The mice were intraperitoneally administered a dose at 250 mg/kg body weight of 5-ALA hydrochloride (Wako Pure Chemical Industries, Ltd., Osaka, Japan) 6–8 weeks after tumor inoculation. Six hours after administration, the excised subcutaneous tumors on the backs of the mice were cut in half. The cut surfaces were examined using fluorescence microscopy and spectroscopy. For HPLC analysis, specimens were collected after necrotic cores and peripheral viable tumors were macroscopically judged by the appearance of the tumors cut in half.
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