Mab1455

Manufactured by R&D Systems

Sourced in United States

MAB1455 is an antibody product offered by R&D Systems. It is a mouse monoclonal antibody that recognizes and binds to an undisclosed target. The antibody is supplied as a lyophilized powder.

Automatically generated - may contain errors

Lab products found in correlation

Mab1368, by R&D Systems (2 mentions) Horse serum, by Thermo Fisher Scientific (1 mentions) Af1924, by R&D Systems (1 mentions) Hoechst, by Thermo Fisher Scientific (1 mentions) Hnf4α, by Santa Cruz Biotechnology (1 mentions) Sc 8987, by Santa Cruz Biotechnology (1 mentions) Live dead kit, by Thermo Fisher Scientific (1 mentions) Af3770, by R&D Systems (1 mentions) Mab36641, by R&D Systems (1 mentions) Alexa fluor 488 and 594, by Jackson ImmunoResearch (1 mentions) Superfrost plus slide, by Thermo Fisher Scientific (1 mentions) Ab150113, by Abcam (1 mentions) Hpa001893, by Merck Group (1 mentions) Cellsens standard 1, by Olympus (1 mentions) Ma5 12023, by Thermo Fisher Scientific (1 mentions) Alexa fluor 568 donkey anti rabbit igg, by Thermo Fisher Scientific (1 mentions) Dp72 microscope, by Olympus (1 mentions) Mouse anti albumin, by R&D Systems (1 mentions) Goat anti rabbit 546 a11010, by Thermo Fisher Scientific (1 mentions) Hoechst 33342, by Merck Group (1 mentions)

10 protocols using mab1455

Immunofluorescence and Viability Assay for Hepatocyte-like Cells

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Immunofluorescence on endoderm and hepatocyte-like cells was performed after fixing cells with 4% PFA for 15 min. Definitive endoderm was marked with antibody SOX-17 1:100 (AF1924, R&D) incubated in 5% horse serum (Thermo Fisher), PBS/Triton X-100 0.3% for 1 h at room temperature (RT). Hepatic endoderm was marked with α-fetoprotein 1:250 (A8452, Sigma-Aldrich). Immature hepatocytes were stained with HNF-4α 1:50 (sc-8987, Santa Cruz Biotechnology). Mature hepatocyte-like cells were stained with albumin 1:100 (MAB1455, R&D), and CYP3A4 (GTX60577, Genetex) incubated in 10% FBS, PBS/Triton X-100 0.1% overnight at 4 °C. For secondary antibody and staining we used Alexa Fluor 594 or 488, phalloidin-488, Hoechst (all Thermo Fisher) for 1 h at 37 °C.
Cell viability assay (Live/Dead kit, Thermo Fisher) was performed on microfluidic differentiated cells. Cells were washed with basal DMEM-F12, incubated with 3 μM ethidium homodimer-1 (stains dead cells in red), 3 μM calcein AM (stains live cells in green) and 4 μM Hoechst (stains cell nuclei in blue) for 45 min at 37 °C, washed with DMEM-F12 and analyzed at fluorescence microscope.

Tonon F., Giobbe G.G., Zambon A., Luni C., Gagliano O., Floreani A., Grassi G, & Elvassore N. (2019). In vitro metabolic zonation through oxygen gradient on a chip. Scientific Reports, 9, 13557.

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Fetal Tissue Immunohistochemistry Protocol

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Fetal tissues were fixed in formalin and embedded in paraffin. Sections of 4–5 μm thickness were cut and placed on Superfrost Plus slides (12–550-17, FisherBrand). For Immunohistochemistry, sections were subjected to heat mediated antigen retrieval (pH6.0) followed by blocking with normal serum. Primary antibodies were incubated overnight at 4°C. The primary antibody we used: GYPA (R&D, MAB1228, 1:250), CD34 (R&D, AF7227, 1:250), CD34 (Novus, NBP2–32933, 1:250), ANXA1 (R&D, AF3770, 1:500), TNFRS10C (R&D, MAB6301, 1:500), AFP (Novus, NBP1–76275, 1:400), ALB (R&D, MAB1455, 1:10K), AHSG (R&D, AF1184, 1:400), and APOA1 (R&D, MAB36641, 1:250). Species and subtype-appropriate fluorescent dye-labelled secondary antibodies were used (Alexa Fluor 488 and 594, 1:400, Jackson ImmunoResearch Lab) or biotinylated secondary antibody were used followed by ABC Elite Systems (PK-6100, Vector Lab) for DAB chromogen staining.

Cao J., O’Day D.R., Pliner H.A., Kingsley P.D., Deng M., Daza R.M., Zager M.A., Aldinger K.A., Blecher R., Zhang F., Spielmann M., Palis J., Doherty D., Steemers F.J., Glass I.A., Trapnell C, & Shendure J. (2020). A human cell atlas of fetal gene expression. Science (New York, N.Y.), 370(6518), eaba7721.

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Immunocytochemistry Evaluation of Hepatic Markers

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Immunocytochemistry evaluated the specific hepatic markers after the induction of differentiation. According to the protocol, the HLC-derived cells were rinsed with PBS and fixed with 4% paraformaldehyde (Sigma-Aldrich, 30525-89-4) for 15 min at 4°C, followed by 5 min at room temperature. After the fixative removal, cells were washed with PBS three times, and Triton X-100 was used to permeabilize cells in PBS. They were incubated with primary mouse monoclonal antibodies with a 1 : 200 dilution against α-fetoprotein (MAB1368; R&D) and albumin (MAB1455; R&D) overnight at 4°C. Next, the cells were rinsed with PBS-Tween 20 (0.1%) three times. They were also incubated with a secondary antibody, anti-goat mouse (1 : 100; F0102B; R&D), and phycoerythrin-conjugated [24 (link)] properly in the darkroom at 37°C for 60 min. Then, these matured cells were rinsed with PBS, and the nuclei eventually were counterstained with 4′, 6-diamidino-2-phenylindole ((DAPI) (1 μg/mL)) (Sigma-Aldrich, 28718-90-3) for 1 min. A fluorescence microscope (Nikon; Japan) was utilized to take images.

Mobarra N., Raji S., Najafi S., Kafi F.K., Ferns G.A, & Pakzad R. (2021). Hypoxia-Induced miR-210 Overexpression Promotes the Differentiation of Human-Induced Pluripotent Stem Cells to Hepatocyte-Like Cells on Random Nanofiber Poly-L-Lactic Acid/Poly (ε-Caprolactone) Scaffolds. Oxidative Medicine and Cellular Longevity, 2021, 4229721.

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Immunofluorescence Staining Protocol

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The cells were fixed using 3.7 % formaldehyde for 10 minutes, and permeablized with 0.1 % triton X-100 for 10 minutes. The cells were incubated with the primary antibody (R&D MAB1455) overnight at 4 °C and then incubated with secondary antibody (abcam ab150113) for 1 hour and washed with phosphate-buffered saline (PBS).

Wu H.H., Ho J.H, & Lee O.K. (2016). Detection of hepatic maturation by Raman spectroscopy in mesenchymal stromal cells undergoing hepatic differentiation. Stem Cell Research & Therapy, 7, 6.

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Immunofluorescence Analysis of SIX1, EYA1, and Cell Markers

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Frozen sections were fixed with 100% ethanol for 15 minutes and permeabilized using 0.05% Tween20 twice (2 minutes per treatment). Blocking was performed using 3% bovine serum albumin for 30 minutes at RT. After washing with phosphate belanced solution (PBS), sections were incubated with rabbit anti-human SIX1 primary antibody (1:100 dilution) (HPA001893, Sigma, St. Louis, MO, USA), rabbit anti-human EYA1 primary antibody (1:100 dilution) (ab85009, Abcam, Cambridge, MA, USA), and mouse anti-human HSA primary antibody (1:1,000 dilution) (MAB1455, R&D Systems, Minneapolis MN, USA) or mouse anti-human GFAP primary antibody (1:100 dilution) (MA5-12023, ThermoFisher, Waltham, MA, USA) at 4 °C overnight. Sections were then incubated with Alexa Fluor^®488 donkey anti-mouse immunoglobulin G (IgG) and Alexa Fluor^®568 donkey anti-rabbit IgG (1:1,000 dilution) (A21202, A10042, ThermoFisher, Waltham, MA, USA) for one hour at RT. After washing with PBS, sections were counterstained with 4',6-diamidino-2-phenylindole (DAPI). Finally, coverslips were mounted with anti-fade mounting medium, and immunofluorescent signals were visualized and recorded using an Olympus DP72 microscope and the cellSens Standard 1.5 software. (Olympus Corporation, Tokyo, Japan)

Xu B., Yang Q., Tang Y., Tan Z., Fu H., Peng J., Xiang X., Gan L., Deng G., Mao Q., Xu P.X., Jiang Y, & Ding J. (2021). SIX1/EYA1 are novel liver damage biomarkers in chronic hepatitis B and other liver diseases. Annals of Translational Medicine, 9(12), 992.

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Antibody Immunodetection in Tissue Samples

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Primary antibodies were used at 0.5–1 μg ml⁻¹ and sourced from Abcam: rabbit anti-Ki67 (ab16667, 1:200), and rabbit anti-pSmad3 (1:200); Fisher Scientific: rabbit anti-follistatin (PA519787, 1:200); Santa Cruz Biotechnologies: anti-Sox2 (sc-17320 1:400); R&D Systems mouse anti-Albumin (MAB1455, 1:1000), rabbit anti-myostatin (AF788, 1:200). Mouse hybridoma anti-eMHC was prepared in house (F1.652 clone, Developmental Studies Hybridoma Bank, University of Iowa, deposited by Blau, HM) and used at 1:50., Secondary fluorochrome conjugated antibodies were from Life Technologies: goat anti-rabbit 546 (A11010, 1:2,000) and goat anti-mouse 488 (A11029, 1:2,000). DNA was stained by Hoechst 33342 from Sigma Aldrich (B2261) used at 1 μg ml⁻¹. Negative controls with isotype-matched immunoglobulin G (IgGs) were routinely performed for all immunodetection studies shown here and above; and background non-specific immunofluorescence was minimal, Supplementary Fig. 8.

Rebo J., Mehdipour M., Gathwala R., Causey K., Liu Y., Conboy M.J, & Conboy I.M. (2016). A single heterochronic blood exchange reveals rapid inhibition of multiple tissues by old blood. Nature Communications, 7, 13363.

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Stepwise Hepatocyte Differentiation from iPSCs

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The step-wise method to produce hepatocytes initiates from differentiating human iPSCs into definitive endoderm, which subsequently patterns into anterior endoderm and further developed into hepatic endoderm. The differentiation method was according to [21 (link), 22 ]. Hepatic cells were further matured to produce cells expressing the hepatic specific protein albumin (ALB). Hepatic differentiation efficiency was examined by immunocytochemistry staining with albumin antibody (R&D System, #MAB1455).

Huang C.Y., Li L.H., Hsu W.T., Cheng Y.C., Nicholson M.W., Liu C.L., Ting C.Y., Ko H.W., Syu S.H., Wen C.H., Yan Z., Huang H.P., Su H.L., Chiang P.M., Shen C.N., Chen H.F., Yen B.L., Lu H.E., Hwang S.M., Chiou S.H., Ho H.N., Wu J.Y., Kamp T.J., Wu J.C, & Hsieh P.C. (2020). Copy number variant hotspots in Han Taiwanese population induced pluripotent stem cell lines - lessons from establishing the Taiwan human disease iPSC Consortium Bank. Journal of Biomedical Science, 27, 92.

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Hepatic Differentiation Marker Expression

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Human ALB and AFP can be produced by mature hepatocytes; thus their antibodies were used to detect the expressions of human ALB and AFP in hADSCs and hUCMSCs during hepatic differentiation. hADSCs and hUCMSCs were inoculated in 24-well plates at a density of 1×10
⁴ cells/mL. On the 3
^rd, 7
^th, 11
^th and 15
^th day after induction, the cells were fixed with 4% formaldehyde, permeabilized with Triton X 100, and blocked with 1% BSA. Then, 1 μL of human serum albumin antibody (MAB1455; R&D) or human alpha fetal protein antibody (MAB13691; R&D) was added to each well. After incubation at 4°C overnight, 1 μL of NL557-conjugated donkey anti-mouse IgG secondary antibody (NL007; R&D) was added and incubated in the dark. After extensive wash with PBS, the expressions of the ALB and AFP proteins in the cells was observed with a Ti-S Eclipse microscope (Nikon, Tokyo, Japan).

Jin Y., Shi R., Qi T., Li Q., Chen C., Gao S., Gao F., Yang D., Sun G., Xu J., Fu Q., Xu J, & Zhang X. (2023). Adipose-derived stem cells show hepatic differentiation potential and therapeutic effect in rats with acute liver failure: Adipose-derived stem cells for acute liver failure. Acta Biochimica et Biophysica Sinica, 55(4), 601-612.

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Immunofluorescence Analysis of Cardiac Markers

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Immunofluorescence analysis was carried out according to standard procedures using secondary Alexa-488 or Alexa-568-conjugated antibodies and Hoechst for fluorescent staining. Primary antibodies were α-actinin (Sigma #A7811, 1:800), albumin (R&D #MAB1455, 1:100, albumin-free staining procedure), ANP (R&D #AF3366, 1:100), α-fetoprotein (Sigma A8452, 1:500), COL3A1 (Santa Cruz #sc-8780-R, 1:100), HA tag (Santa Cruz #sc-806-X, 1:200), MLC2v (ProteinTech Group #10906-1-AP, 1:200), NKX2.5 (R&D #AF2444, 1:100), PECAM1 (R&D #BBA7, 1:50), smooth muscle actin (DakoCytomation #M085129-2, 1:100), SOX2 (R&D #AF2018, 1:200), cardiac troponin I (cTnI, Santa Cruz #sc-15368, 1:200), cardiac troponin T (cTnT, Thermo #MS-295-P, 1:200), renin (R&D #AF4090, 1:150), β-III-tubulin (Covance #MMS-435P, 1:500), and vimentin (Sigma #V6630, 1:200).

Pfeiffer M.J., Quaranta R., Piccini I., Fell J., Rao J., Röpke A., Seebohm G, & Greber B. (2018). Cardiogenic programming of human pluripotent stem cells by dose-controlled activation of EOMES. Nature Communications, 9, 440.

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Immunofluorescence Analysis of Hepatocyte-like Cells

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After completing the differentiating process, the hHLCs were rinsed twice with PBS and fixed with 4% paraformaldehyde (Sigma, 30525-89-4) for 20 min at 4°C, then 5 min at room temperature (RT). After washing, the cells were permeabilized with 3% Triton X-100 in PBS.
The fixed cells were blocked for 30 min at 37°C with 10% goat serum (Gibco, 16210064)/PBS and then remove it. They were next incubated overnight at 4°C in a humidity chamber with respective primary antibodies: mouse monoclonal anti-fetoprotein (AFP, 1:200, R&D, MAB1368), mouse monoclonal anti-albumin (ALB, 1: 200, R&D, MAB1455). At the end of the incubation time, the cells were rinsed 3 times with PBS-Tween 20 (0.1%) and incubated with the PE conjugated as a secondary antibody, goat anti-mouse (1:100; R&D, F0102B) as appropriate for 45 min at 37 °C in the dark. Subsequently, these subjected cells were washed with PBS-Tween %0.1, 3×5 min. The nuclei were counterstained with 4′, 6-diamidino-2-phenylindole [(DAPI) (1 μg/ml)] (Sigma, 28718-90-3). Nuclear staining and the cells were analyzed with a fluorescent microscope (Nikon; Japan).

Mobarra N., Soleimani M., Kouhkan F., Hesari Z., Lahmy R., Mossahebi-Mohammadi M., Arefian E., Jaafarpour Z., Nasiri H., Pakzad R., Tavakoli R, & Pasalar P. (2014). Efficient Differentiation of Human Induced Pluripotent Stem Cell (hiPSC) Derived Hepatocyte-Like Cells on hMSCs Feeder. International Journal of Hematology-Oncology and Stem Cell Research, 8(4), 20-29.

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