Nitrocellulose

Manufactured by Pall Corporation

Sourced in New Zealand, United States

Nitrocellulose is a type of lab equipment used in various scientific applications. It is a membrane material made from cellulose that has been chemically treated with nitric acid. Nitrocellulose membranes are commonly used for protein and nucleic acid transfer and detection in analytical techniques such as Western blotting and Northern blotting.

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Lab products found in correlation

Image studio 4, by LI COR (2 mentions) Odyssey clx infrared imaging system, by LI COR (2 mentions) Mini beadbeater, by Biospec (1 mentions) Gel image analysis system, by Bio-Rad (1 mentions) Gradient sds polyacrylamide gel, by Bio-Rad (1 mentions) Mini protean tgx precast protein gel, by Bio-Rad (1 mentions) Protease arrest, by G Biosciences (1 mentions) Blocker casein, by Thermo Fisher Scientific (1 mentions) Cell lysis buffer, by Cell Signaling Technology (1 mentions) Rabbit anti iκb, by Cell Signaling Technology (1 mentions) Rabbit anti actin, by Cell Signaling Technology (1 mentions) Pierce bca protein assay kit, by Thermo Fisher Scientific (1 mentions) Polyvinylidene fluoride (pvdf), by Merck Group (1 mentions) Alliance mini chemiluminescence device, by Uvitec (1 mentions) Chemiluminescence assay kit, by Merck Group (1 mentions)

Spelling variants of this product

Nitrocellulose membranes (251 citations) Nitrocellulose filter membrane (16 citations) Nitrocellulose blotting membranes (9 citations) Nitrocellulose (NC) membrane (9 citations) BioTrace NT Nitrocellulose Transfer Membrane (6 citations) 0.2 μm nitrocellulose membranes (4 citations) BioTrace NT pure nitrocellulose blotting membrane (3 citations) Pure nitrocellulose (2 citations) NT nitrocellulose membranes (2 citations) 0.45 μm nitrocellulose membrane (2 citations)

5 protocols using nitrocellulose

Protein Extraction and Analysis from Microalgae

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Samples were collected from cultures in the late exponential phase. Cells were disrupted with a Mini Bead Beater (Biospec Products, Oklahoma) at 3500 rpm for 20 s in the presence of glass beads (150–212 mm diameter), B1 buffer (400 mM NaCl, 2 mM MgCl₂, and 20 mM Tricine–KOH, pH 7.8), 0.5% milk powder, 1 mM PMSF, 1 mM DNP‐e‐amino‐n‐caproic acid, and 1 mM benzamidine. The ruptured cells were then solubilized in a buffer (×3) contained 30% glycerol, 125 mM Tris, pH 6.8, 0.1 M dithiothreitol, and 9% SDS at RT for 20 min. SDS‐PAGE analysis was performed with a Tris‐glycine buffer system as previously described (Laemmli, 1970 (link)) with acrylamide at a final concentration of 12%. Western blot analysis was performed by transferring the proteins to nitrocellulose (Bio Trace, Pall Corporation, Auckland, New Zealand) and detecting them with alkaline phosphatase‐conjugated antibodies. The antibodies recognized the PSI subunits PsaA and PsaD (Agrisera), D2, LHCX1, VCP, and LHCII proteins (antibodies produced by immunizing New Zealand rabbits with purified spinach protein [D2, LHCII] or recombinant N. gaditana proteins [VCP and LHCX1]; Meneghesso et al., 2016 (link)).

Fattore N., Savio S., Vera‐Vives A.M., Battistuzzi M., Moro I., La Rocca N, & Morosinotto T. (2021). Acclimation of photosynthetic apparatus in the mesophilic red alga Dixoniella giordanoi. Physiologia Plantarum, 173(3), 805-817.

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Evaluating Protein Signaling Pathways

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After incubation with Cd, cells were washed twice with PBS and lysed in radioimmunoprecipitation assay (RIPA) lysis buffer for 30 min at 4℃. Following sonication and centrifugation, the protein concentration of the cells was measured using a BCA protein assay kit according to the manufacturer's instructions. The protein samples were separated using 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), transferred to nitrocellulose (Pall Corporation, USA), and then incubated overnight with the primary antibodies (rabbit anti-rat antibodies to P-p38, p38, P-ERK, ERK, P-JNK, JNK, cleaved PARP, cleaved caspase-9, cleaved caspase-3, Bax, and Bcl-2 at dilutions of 1 : 1000; anti-β-actin antibody was used at a dilution of 1 : 5000) at 4℃, followed by incubation with HRP-conjugated goat anti-rabbit IgG for 2 h at 1 : 5000. Proteins were visualized using enhanced chemiluminescence detection reagents. The band intensity was determined by a gel image analysis system (Bio-Rad Laboratories, USA) and normalized with β-actin.

Zhao H., Liu W., Wang Y., Dai N., Gu J., Yuan Y., Liu X., Bian J, & Liu Z.P. (2015). Cadmium induces apoptosis in primary rat osteoblasts through caspase and mitogen-activated protein kinase pathways. Journal of Veterinary Science, 16(3), 297-306.

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Quantitative Western Blotting Technique

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Western blotting was performed using 30–50 μg total cellular protein per sample. Protein was loaded and separated over 4–20% gradient SDS polyacrylamide gels (Bio-Rad, Hercules, CA). Proteins were transferred to nitrocellulose (Pall Corporation, Ann Arbor, MI), and blocked for 60 min in 0.1% casein in TBS. Blots were incubated in 0.1% casein/TBS-T with a 1:1000 dilution of primary antibody overnight at 4°C with gentle rocking. Blots were washed six times for 5 min each in TBS-T, and then incubated with infrared-labeled secondary antibodies at 1:15,000 final dilution. in Blocker^™ Casein in TBS with 0.1% TWEEN^® 20 and incubated at room temperature for 1 hour with gentle rocking. Blots were visualized and quantified using a LiCor Odyssey CLx Infrared imaging system (Lincoln, NE). After background subtraction, fluorescence intensity for the protein of interest was normalized to the signal intensity for the relevant loading control, using Image Studio 4.0 (Li-Cor, Lincoln, Nebraska).

Ndaw V.S., Abebayehu D., Spence A.J., Paez P.A., Kolawole E.M., Taruselli M.T., Caslin H.L., Chumanevich A.P., Paranjape A., Baker B., Barnstein B.O., Haque T.T., Kiwanuka K.N., Oskeritzian C.A, & Ryan J.J. (2017). TGFβ1 Suppresses IL-33-induced Mast Cell Function. Journal of immunology (Baltimore, Md. : 1950), 199(3), 866-873.

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Western Blot Analysis of IκB Protein

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Cell lysates were collected using Protease arrest (GBiosciences, Maryland Heights, MO) in cell lysis buffer (Cell Signaling Technology, Danvers, MA). Protein concentration was determined using the Pierce BCA Protein Assay Kit (Thermofisher, Waltham, MA). 4–20% Mini-PROTEAN® TGX™ Precast Protein Gels (Bio–Rad, Hercules, CA) were loaded with 30 μg protein, electrophoresed and transferred to nitrocellulose (Pall Corporation, Ann Arbor, MI), and membranes were blocked for 60 minutes in Blocker casein in tris-buffered saline (TBS) (from Thermofisher, Waltham, MA). Blots were incubated with primary antibodies overnight in block buffer + Tween20 (1:1000) with rabbit anti-IκB and rabbit anti-actin (1:1000, antibodies all purchased from Cell Signaling, Danvers, MA). Blots were washed six times for 5 minutes each in TBS-Tween-20, followed by incubation with secondary antibody (1:10,000) for 60 minutes at room temperature (Cell Signaling, Danvers, MA). Size estimates for proteins were obtained using molecular weight standards from Bio–Rad (Hercules, CA). Blots were visualized and quantified using a LiCor Odyssey CLx Infrared imaging system (Lincoln, NE). After background subtraction, fluorescence intensity for the protein of interest was normalized to the signal intensity for the actin calculated relative to unactivated samples, using Image Studio 4.0 (LiCor).

Caslin H.L., Abebayehu D., Qayum A.A., Haque T.T., Taruselli M.T., Paez P.A., Pondicherry N., Barnstein B.O., Hoeferlin L.A., Chalfant C.E, & Ryan J.J. (2019). Lactic acid inhibits LPS-induced mast cell function by limiting glycolysis and ATP availability. Journal of immunology (Baltimore, Md. : 1950), 203(2), 453-464.

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Western Blot Analysis of STAT3 Phosphorylation

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BMDM lysates were collected in Protein 5× Sample Buffer (ELPIS BIOTECH, EBA-1052) diluted with RIPA buffer (150 mM sodium chloride, 1% Triton X-100, 0.1% SDS, 1% sodium deoxycholate, 50 mM Tris-Cl at pH 7.5, and 2 mM EDTA) supplemented with protease (04693132001) and phosphatase (11836170001) inhibitor cocktails (Roche, Mannheim, Germany). Samples were boiled for 10 min on a heating block and cooled on ice for 10 min. The samples were resolved by SDS-PAGE and transferred to a nitrocellulose (Pall Corporation, NY, 66485) or PVDF (Millipore, Burlington, MA, IPVH0001) membrane at 200 mA for 2 h. To prevent nonspecific binding, membranes were incubated in blocking solution with 1% BSA in TBST for 30 min at room temperature and reacted overnight with anti-pSTAT3, -STAT3, or -ACTIN primary antibody at 4°C. The membranes were reacted with the appropriate horseradish peroxidase-conjugated secondary antibodies for 1 h at room temperature. Immunoreactive bands were visualized with ECL reagent from the Chemiluminescence Assay Kit (Millipore, WBKL S0500), and the appropriate bands were detected using the UVitec Alliance mini-chemiluminescence device (UVitec, Rugby, UK). Band intensities were measured using Image J software and normalized to that of ACTIN.

Kim Y.J., Park E.J., Lee S.H., Silwal P., Kim J.K., Yang J.S., Whang J., Jang J., Kim J.M, & Jo E.K. (2023). Dimethyl itaconate is effective in host-directed antimicrobial responses against mycobacterial infections through multifaceted innate immune pathways. Cell & Bioscience, 13, 49.

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