Tlc plate

Manufactured by Merck Group

Sourced in Germany, United States, India

TLC plates are a type of laboratory equipment used for thin-layer chromatography, a separation technique used to analyze and purify chemical compounds. TLC plates are typically made of glass, aluminum, or plastic and coated with a thin layer of adsorbent material, such as silica gel or alumina. These plates allow for the separation and identification of components in a mixture by their differential migration on the adsorbent surface when a solvent is applied.

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Lab products found in correlation

Silica gel, by Merck Group (6 mentions) Silica gel 60, by Merck Group (6 mentions) γ 32p atp, by PerkinElmer (5 mentions) Bca protein assay kit, by Thermo Fisher Scientific (4 mentions) Silica gel 60 f254, by Merck Group (4 mentions) Dansyl chloride, by Merck Group (3 mentions) Proline, by Merck Group (3 mentions) Toluene, by Merck Group (3 mentions) Quercetin, by Merck Group (3 mentions) Dimethyl sulfoxide (dmso), by Merck Group (3 mentions) Acetic acid, by Merck Group (3 mentions) Storage phosphor screen, by GE Healthcare (3 mentions) Methanol, by Merck Group (3 mentions) Typhoon fla 7000, by GE Healthcare (3 mentions) Bas ms 2340, by Fujifilm (2 mentions) Ascorbic acid, by Merck Group (2 mentions) Whatman no 1 filter paper, by Cytiva (2 mentions) Dl dap, by Merck Group (2 mentions) Petroleum ether, by Merck Group (2 mentions) Diethyl ether, by Merck Group (2 mentions)

177 protocols using tlc plate

Enzymatic Oligosaccharide Digestion and Analysis

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All oligosaccharide substrates and standards for digests and TLC analyses were purchased from Megazyme, except for (GlcN)₂, which was purchased from Sigma. Digest reactions were carried out in McIlvaine phosphate–citrate buffer pH 6.5 using an oligosaccharide substrate concentration of 10 mg ml⁻¹ and 0.5 µM VC0615. 20 µl reaction volumes containing enzyme and substrate were incubated at 37°C with shaking. At set time points, 2 µl of the reaction mixture was removed and ‘paused’ by flash-freezing in liquid N₂. Following completion of the reaction time course, all frozen aliquots were thawed and spotted onto an aluminium foil-backed silica TLC plate (Sigma).
The TLC plate was run in a 50:25:25 n-butanol:water:acetic acid solvent system until the solvent front reached ∼1–2 cm from the top of the plate. In order to improve the separation between spots, the TLC plate was dried and rerun a second time in the same solvent system. Following the second run, the TLC plate was dried and developed using a p-anisaldehyde (Sigma) stain (3.7 ml p-anisaldehyde, 1.5 ml acetic acid, 5 ml concentrated sulfuric acid, 135 ml ethanol) with mild heating.

Wu L, & Davies G.J. (2018). Structure of the GH9 glucosidase/glucosaminidase from Vibrio cholerae. Acta Crystallographica. Section F, Structural Biology Communications, 74(Pt 8), 512-523.

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Extraction and Purification of Siderophore

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By using a silica gel thin-layer chromatography (TLC) plate (Merck-Germany) and a Dichloromethane (DCM): Methanol (9:1 v/v) as a solvent system, the siderophore compound was extracted as follows: in a chromatographic jar was initially equilibrated with the solvent system for 20 min, the TLC plate that loaded with ethyl acetate extract was subsequently immersed in the solvent system to facilitate complete elution and separation of compounds. Then, the TLC plate was carefully removed and visualized under a UV lamp (VL-215 LC, Marne La Vallee, France) at 254/365 nm. The desired band that showed a bright blue fluorescence, was carefully scratched, transferred to another vial, and eluted several times using ethyl acetate to obtain the siderophore compound. This process was repeated multiple times to obtain the desired quantity of the compound, which was then purified using a sephadex LH-20 column and a DCM: Methanol (6:4 v/v) solvent system.

Khazaal M.T., Faraag A.H.I., Hamada M.A, & El-Hendawy H.H. (2024). Characterization and Statistical Optimization of Enterobatin Synthesized by Escherichia coli OQ866153. Biochemical genetics.

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Determination of Actinomycetes Genus

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In order to determine the genus of antagonistic actinomycetes, the 2,6- diaminopimelic acid, one of the cell wall components of actinomycetes mycelia was analyzed using the method of Hasegawa et al. (1983) [38 (link)]. The selected strain was cultured in ISP-2 medium (4.0 g yeast extract, 10.0 g malt extract, 4.0 g dextrose, and 1 L sterile distilled water, pH 7.3) at 150 rpm, 28℃ for 14 days. After cultivation, the culture broth was centrifuged to collect cells. 6N HCl was used to hydrolyze the cells by heating at 70ºC for 18 h in a water bath. The hydrolysate was filtered through Whatman No.1 filter paper and evaporated to dryness in order to remove the HCl residue. Dried hydrolysate was dissolved in 1 mL of distilled water and applied onto TLC plate (15 x 20 cm, Merck Co., USA). A total of 20 ul of 0.01 M DL-DAP (Sigma Chemical Co., USA) containing both meso- and LL-DAP isomers and amino acids (alanine, glycine and glutamate) was also loaded on the TLC plates as a standard.

Chaiharn M., Theantana T, & Pathom-aree W. (2020). Evaluation of Biocontrol Activities of Streptomyces spp. against Rice Blast Disease Fungi. Pathogens, 9(2), 126.

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Purification and Thin-Layer Chromatography of Lipids

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To get rid of unspecifically associated lipids, purified YnaI samples were cleaned up again with the Superose 6 column. TLC experiment was performed as previously described^{35 (link),36 (link)}. Briefly, lipids from the samples were extracted with the mixed solution of chloroform and methanol (2:1). TLC plate (Millipore Sigma) was impregnated with 1.8% boric acid in ethanol, air dried and then activated at 100 °C for 30 min. Samples and lipid standards were dotted on the TLC plate. The experiment was performed with a mobile phase of chloroform-ethanol-water-triethylamine (30:35:7:35, v/v/v/v) for 2–3 h. The plate was stained in 50% ethanol with 3.2% sulfuric acid and 0.5% MnCl₂ for 20 s, air dried, baked at 120 °C for 30 min, and imaged for analysis.

Hu W., Wang Z, & Zheng H. (2021). Mechanosensitive channel YnaI has lipid-bound extended sensor paddles. Communications Biology, 4, 602.

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Purification and Lipid Analysis of SLC26A2

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SLC26A2 samples were subjected to two rounds of purification using the Superose 6 column to remove non-specifically associated lipids. The TLC experiment was performed following a previously described method^{60 (link)}. Specifically, the lipids tightly associated with SLC26A2 were extracted using a mixed solution of chloroform and methanol (2:1). A TLC plate (Millipore Sigma) was washed with ethanol, air dried, and then activated at 100 °C for 30 min. SLC26A2 samples and lipid standards were dotted on the TLC plate. The experiment was performed with a first mobile phase of chloroform-methanol-water (65:25:4, v/v/v) till the solvent front was halfway through the plate, followed by a second mobile phase of hexane-acetone (100:1, v/v) till the solvent front reached the top. After the run, the TLC plate was air dried, stained with 0.03% (w/v) G-250 in 20% methanol for 15 min, destained in 20% methanol for 10 min, and then air dried again.

Hu W., Song A, & Zheng H. (2024). Substrate binding plasticity revealed by Cryo-EM structures of SLC26A2. Nature Communications, 15, 3616.

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Decapping Assay for Capped RNA

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The decapping assay was performed by incubating 2μM of recombinant DXO with 1μM of capped RNA in IVDA-2 buffer (10mM Tris-HCl pH 7.5, 100mM KOAc, 2mM MgCl₂, 0.5mM MnCl₂, 2mM DTT, 0.2mM spermidine) at 37°C for 2 hours. Alternatively, each RNA substrate was treated with 0.5U NucP1 (Sigma-Aldrich, #N8630) in 100mM NaOAc pH 5.2. The reaction was stopped by adding formic acid to a final concentration of 500mM. Aliquots of the reactions (5CPM in lanes 1 and 3–12, 10CPM in lane 2) were applied to a polyethyleneimine-cellulose TLC plate (Sigma-Aldrich, #Z122882), which was developed with 0.3M (NH₄)₂SO₄. The release of different cap products was revealed by autoradiography with a phosphorimager (Amersham Biosciences).

Picard-Jean F., Brand C., Tremblay-Létourneau M., Allaire A., Beaudoin M.C., Boudreault S., Duval C., Rainville-Sirois J., Robert F., Pelletier J., Geiss B.J, & Bisaillon M. (2018). 2'-O-methylation of the mRNA cap protects RNAs from decapping and degradation by DXO. PLoS ONE, 13(3), e0193804.

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ATPase Activity Assay of PfPSH1C

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To analyze the ATPase activity, the release of inorganic phosphate (Pi) from [γ‐³²P] ATP by the action of PfPSH1C was measured. To perform standard ATPase assay reaction, purified PfPSH1C protein was used and the buffer used for the assay contains 20 mm Tris/HCl, pH 8.0, 8 mm DTT, 1.0 mm MgCl₂, 20 mm KCl, 16 μg·ml⁻¹ BSA, and 50 ng of M13 mp19 ssDNA. A mixture of 1 μL [γ‐³²P]‐labeled ATP (~ 17 nm) and 1 mm cold ATP was added later to the reaction. The reaction was incubated for 1 h at 37 °C, and the reaction was quenched on ice. One microlitre of the quenched reaction was spotted on TLC plate (Sigma), and the hydrolyzed Pi was separated on TLC using TLC buffer (0.5 m LiCl and 1 m formic acid). The TLC plate was exposed to the phosphoimager film and then scanned on a phosphoimager, and the raw files were converted to TIFF images and processed using imagej software (NIH, Bethesda, MD, USA).

Chauhan M., Sourabh S., Yasmin R., Pahuja I, & Tuteja R. (2019). Biochemical characterization of Plasmodium falciparum parasite specific helicase 1 (PfPSH1). FEBS Open Bio, 9(11), 1909-1927.

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Dansylation and TLC of Polyamines

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Polyamines were separated by thin-layer chromatography as previously described [30 (link)]. For all samples, cells were treated as described prior to being trypsinized and centrifuged. Pellets were washed with PBS and then resuspended in 200 µL 2% perchloric acid. Samples were then incubated overnight at 4 °C. 200 µL of supernatant was combined with 200 µL 5 mg/mL dansyl chloride (Sigma Aldrich) in acetone and 100 µL saturated sodium bicarbonate. Samples were incubated in the dark overnight at room temperature. Excess dansyl chloride was cleared by incubating the reaction with 100 µL 150 mg/mL proline (Sigma Aldrich). Dansylated polyamines were extracted with 50 µL toluene (Sigma Aldrich) and centrifuged. Five microliter of sample was added in small spots to the TLC plate (silica gel matrix; Sigma Aldrich) and exposed to ascending chromatography with 1:1 cyclohexane: ethyl acetate. The plate was dried and visualized via exposure to UV.

Dial C.N., Tate P.M., Kicmal T.M, & Mounce B.C. (2019). Coxsackievirus B3 Responds to Polyamine Depletion via Enhancement of 2A and 3C Protease Activity. Viruses, 11(5), 403.

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ATPase Activity Assay of PfPSH3N Protein

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The ATPase activity was assayed by measuring the released Pi from [γ-³²P] ATP. The purified PfPSH3N protein was mixed with buffer (20 mM Tris–HCl, pH 8.0, 8 mM DTT, 1.0 mM MgCl₂, 20 mM KCl and 16 μg/ml BSA) and a mixture of [γ-³²P] ATP (~17 nM) and 1 mM cold ATP and it was incubated at 37 °C for 1 hr. For checking DNA-dependant ATPase activity 50 ng of M13 mp19 ssDNA was added in the above reaction and negative control was without protein. The reaction was quenched on ice after 1 hr of incubation and then 1 μl of reaction mix was spotted onto thin layer chromatography (TLC) plate (Sigma). TLC buffer (0.5 MLiCl and 1 M formic acid) was used for separating hydrolysed Pi using TLC and the plate was air dried and then scanned on phosphoimager. All the quantitation was done using Image j software^{61 (link)}.

Chauhan M., Tarique M, & Tuteja R. (2017). Plasmodium falciparum specific helicase 3 is nucleocytoplasmic protein and unwinds DNA duplex in 3′ to 5′ direction. Scientific Reports, 7, 13146.

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Thin Layer Chromatography of Lipids

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TLC was performed according to Yao and Rastetter^{50 (link)}. A TLC plate (silica on aluminum, Sigma) was washed with methanol and dried for 30 min at 150 °C. A full length predevelopment was performed in methyl acetate:1-propanol:chloroform:methanol:0.25%KCl (25:25:25:10:9, v/v/v/v/v) followed by drying for 30 min. 1.5 µg lipid per lane was applied onto the TLC plate. The TLC plate was developed halfway using the solvent used for predevelopment. The plate was dried and developed until the solvent front was about 1 cm beyond the end of the plate using hexane:diethyl ether:acetic acid (75:23:2). The plate was dried for another 30 min and finally totally developed using hexane. Detection was done by applying 10% copper sulfate hydrate in 8% phosphoric acid, followed by heating at 200 °C. Standard lipids were purchased from Lipoid.

Vorselen D., van Dommelen S.M., Sorkin R., Piontek M.C., Schiller J., Döpp S.T., Kooijmans S.A., van Oirschot B.A., Versluijs B.A., Bierings M.B., van Wijk R., Schiffelers R.M., Wuite G.J, & Roos W.H. (2018). The fluid membrane determines mechanics of erythrocyte extracellular vesicles and is softened in hereditary spherocytosis. Nature Communications, 9, 4960.

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