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Meridian Life Science polymerase chain reaction pcr mixtures
Polymerase Chain Reaction Pcr Mixtures, supplied by Meridian Life Science, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 91 stars, based on 1 article reviews
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polymerase chain reaction pcr mixtures - by Bioz Stars, 2020-09
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Article Title: The fungal composition of natural biofinishes on oil-treated wood
Article Snippet: .. The polymerase chain reaction (PCR) mixtures had final concentrations of: 4% DNA extract, 10% PCR buffer, 3% MgCl2 (25 mM), 65.8% demineralised sterile water, 7.8% dNTP (1 mM), 5% DMSO, 2% forward primer (10 μM), 2% reverse primer and 0.4% Taq polymerase (5 U/mL, BioTaq, Bioline). .. The PCR program typically consisted of 1 cycle of 5 min denaturation at 95 °C; 35 cycles of 35 s denaturation at 95 °C, followed by ITS-primer annealing for 30 s at 55 °C or LSU -primer annealing at 54 °C for 50 s, and an extension for 1.5 min at 72 °C.

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    Meridian Life Science psa
    AR stimulates PP1α recruitment and mobilization of P-TEFb. ( A and B ) LNCaP cells in CDS medium were treated for 2 h (A) or 2–8 h (B) with androgen (10 nM DHT) as indicated, followed by ChIP-qPCR for AR and PP1α binding to the indicted AREs. ( C ) LNCaP cells in CDS medium were treated for 4 h with PP1α inhibitor tautomycin (Tau, 200 nM) and DHT, followed by ChIP-qPCR. ( D ) LNCaP in CDS medium were treated overnight with PP2A-specific inhibitor fostriecin (Fos, 100 nM) and DHT, followed by ChIP-qPCR. ( E ) LNCaP cells in CDS medium were treated for 4 h with tautomycin and DHT as indicated, followed by isolation of cytoplasmic, nuclear and chromatin fractions, with HSP90, <t>FOXA1</t> and H3 used as marker proteins for the cellular fractions, respectively (right panel). The chromatin fraction was then immunoblotted for the indicated proteins (left panel). ( F–H ) LNCaP cells in CDS medium were transfected with control siRNA, AR-specific siRNA or two independent PP1α siRNAs (20 nM) for 3 days, followed by treatment with DHT (10 nM). Total proteins were normalized and immunoblotted (F); cells were harvested for ChIP-qPCR analysis (G); or qRT-PCR analysis was carried out for <t>PSA</t> of KLK2 pre-mRNA (values normalized to the basal activity without DHT, set to 1) (H).
    Psa, supplied by Meridian Life Science, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 91 stars, based on 1 article reviews
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    psa - by Bioz Stars, 2020-09
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    92
    Meridian Life Science rotavirus
    Immunohistochemical analysis and viral challenge. Immunof-luorescent analysis of the virus-neutralizing effects of IgG antibodies directed against HRVVP7. (A) Negative control group, (B) HRVVP7-immunized group at 1:42 dilution, (C) HRVVP7-linker-CTB-immunized group at 1:42 dilution and (D) HRVVP7-linker-CTB-immunized group at 1:67 dilution. Magnification, ×125. (E) Incidence of diarrhea in neonatal suckling mice following challenge with simian <t>rotavirus</t> SA11. ***P
    Rotavirus, supplied by Meridian Life Science, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Meridian Life Science human pancreatic α amylase
    Expression of <t>α‐amylase</t> in normal human tissues. The mRNA expressions of α‐amylase in human normal tissues were quantified by real‐time PCR. The α‐amylase/GAPDH ratio for liver was set as the baseline value to which all transcript levels were normalized. mRNA of α‐amylase/GAPDH in human tissues I and II across Human MTC Panel I and II (A) and in human digestive tissues across the Human Digestive System MTC Panel (B). GAPDH, glyceraldehyde‐3‐phosphate dehydrogenase; mRNA, messenger RNA; MTC, multiple tissue cDNA; PCR, Polymerase chain reaction
    Human Pancreatic α Amylase, supplied by Meridian Life Science, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    AR stimulates PP1α recruitment and mobilization of P-TEFb. ( A and B ) LNCaP cells in CDS medium were treated for 2 h (A) or 2–8 h (B) with androgen (10 nM DHT) as indicated, followed by ChIP-qPCR for AR and PP1α binding to the indicted AREs. ( C ) LNCaP cells in CDS medium were treated for 4 h with PP1α inhibitor tautomycin (Tau, 200 nM) and DHT, followed by ChIP-qPCR. ( D ) LNCaP in CDS medium were treated overnight with PP2A-specific inhibitor fostriecin (Fos, 100 nM) and DHT, followed by ChIP-qPCR. ( E ) LNCaP cells in CDS medium were treated for 4 h with tautomycin and DHT as indicated, followed by isolation of cytoplasmic, nuclear and chromatin fractions, with HSP90, FOXA1 and H3 used as marker proteins for the cellular fractions, respectively (right panel). The chromatin fraction was then immunoblotted for the indicated proteins (left panel). ( F–H ) LNCaP cells in CDS medium were transfected with control siRNA, AR-specific siRNA or two independent PP1α siRNAs (20 nM) for 3 days, followed by treatment with DHT (10 nM). Total proteins were normalized and immunoblotted (F); cells were harvested for ChIP-qPCR analysis (G); or qRT-PCR analysis was carried out for PSA of KLK2 pre-mRNA (values normalized to the basal activity without DHT, set to 1) (H).

    Journal: Nucleic Acids Research

    Article Title: Positive feedback loop mediated by protein phosphatase 1α mobilization of P-TEFb and basal CDK1 drives androgen receptor in prostate cancer

    doi: 10.1093/nar/gkw1291

    Figure Lengend Snippet: AR stimulates PP1α recruitment and mobilization of P-TEFb. ( A and B ) LNCaP cells in CDS medium were treated for 2 h (A) or 2–8 h (B) with androgen (10 nM DHT) as indicated, followed by ChIP-qPCR for AR and PP1α binding to the indicted AREs. ( C ) LNCaP cells in CDS medium were treated for 4 h with PP1α inhibitor tautomycin (Tau, 200 nM) and DHT, followed by ChIP-qPCR. ( D ) LNCaP in CDS medium were treated overnight with PP2A-specific inhibitor fostriecin (Fos, 100 nM) and DHT, followed by ChIP-qPCR. ( E ) LNCaP cells in CDS medium were treated for 4 h with tautomycin and DHT as indicated, followed by isolation of cytoplasmic, nuclear and chromatin fractions, with HSP90, FOXA1 and H3 used as marker proteins for the cellular fractions, respectively (right panel). The chromatin fraction was then immunoblotted for the indicated proteins (left panel). ( F–H ) LNCaP cells in CDS medium were transfected with control siRNA, AR-specific siRNA or two independent PP1α siRNAs (20 nM) for 3 days, followed by treatment with DHT (10 nM). Total proteins were normalized and immunoblotted (F); cells were harvested for ChIP-qPCR analysis (G); or qRT-PCR analysis was carried out for PSA of KLK2 pre-mRNA (values normalized to the basal activity without DHT, set to 1) (H).

    Article Snippet: Antibodies and immunoblotting The sources for the antibodies and control IgGs were as following: AR (Santa Cruz, Cat. sc-816); pAR-S81 (EMD Millipore, Cat. 07–1375); pAR-S308 (Santa Cruz, Cat. sc-26406); pRNA Pol II Ser2 (Abcam, Cat., ab5095); pRNA Pol II Ser5 (Abcam, Cat. ab5131); CDK1 (Cell Signaling, Cat. 9112); pCDK1-T161 (Cell Signaling, Cat. 9114); CDK9 (Santa Cruz, Cat. sc-8338); Cyclin T1 (Santa Cruz, Cat. sc-10750); BRD4 (Bethyl, Cat. A301-985A); p300 (Santa Cruz, Cat. sc-585); Histone 3 (Abcam, Cat. ab1791); H3K27Ac (Abcam, Cat. ab4729); pH3-Ser10 (EMD Millipore, Cat. 06–570); FoxA1 (Abcam, Cat. Ab23738); PSA (Meridian Life Science, Cat. K92110R); Flag-M2 (Sigma-Aldrich, Cat. F3165); HA (Cell Signaling, Cat. 3724); β-Tubulin (EMD Millipore, Cat. MAB3408); β-Actin (Abcam, Cat. Ab6276); GAPDH (Abcam, Cat. Ab9485); Hsp90 (Santa Cruz, Cat. sc-69703), PP1 (Santa Cruz: Cat. sc-443; Cat. sc-6104; and Cat. sc-6105), and control IgGs (Santa Cruz: normal rabbit IgG, Cat. sc-2027; and normal goal IgG, Cat. sc-2028).

    Techniques: Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Binding Assay, Isolation, Marker, Transfection, Quantitative RT-PCR, Activity Assay

    Immunohistochemical analysis and viral challenge. Immunof-luorescent analysis of the virus-neutralizing effects of IgG antibodies directed against HRVVP7. (A) Negative control group, (B) HRVVP7-immunized group at 1:42 dilution, (C) HRVVP7-linker-CTB-immunized group at 1:42 dilution and (D) HRVVP7-linker-CTB-immunized group at 1:67 dilution. Magnification, ×125. (E) Incidence of diarrhea in neonatal suckling mice following challenge with simian rotavirus SA11. ***P

    Journal: Experimental and Therapeutic Medicine

    Article Title: Oral immunization with rotavirus VP7-CTB fusion expressed in transgenic Arabidopsis thaliana induces antigen-specific IgA and IgG and passive protection in mice

    doi: 10.3892/etm.2018.6003

    Figure Lengend Snippet: Immunohistochemical analysis and viral challenge. Immunof-luorescent analysis of the virus-neutralizing effects of IgG antibodies directed against HRVVP7. (A) Negative control group, (B) HRVVP7-immunized group at 1:42 dilution, (C) HRVVP7-linker-CTB-immunized group at 1:42 dilution and (D) HRVVP7-linker-CTB-immunized group at 1:67 dilution. Magnification, ×125. (E) Incidence of diarrhea in neonatal suckling mice following challenge with simian rotavirus SA11. ***P

    Article Snippet: The intracellular rotavirus was detected using the mouse mAb directed against rotavirus VP7 (cat. no. C01715M; Meridian Life Science, Inc., Memphis, TN, USA; 1:500 dilution) as the primary antibody at 25°C for 30 min and fluorescein-isothiocyanate-labeled goat anti-mouse IgG antibody (cat. no. ab6785; Abcam) as the secondary antibody (1:500 dilution) at 37°C for 1 h. The cells were washed three times with PBS at room temperature (15 min/wash).

    Techniques: Immunohistochemistry, Negative Control, CtB Assay, Mouse Assay

    Schematic of plant expression plasmids pPHAP1301-HRVVP7-CTB and pPHAP1301-HRVVP7. bar encodes a resistance marker for glufosinate selection. (A) HRVVP7 coding sequence. (Gly 4 Ser) 3 is the linker between HRVVP7 and CTB . (B) HRVVP7 coding sequence. HRVVP7, human rotavirus VP7; CTB, cholera toxin B subunit; Phas , β-phaseolin storage protein; 35s, Cauliflower mosaic virus; LB, left border; RB, right border; nos , nopaline synthase.

    Journal: Experimental and Therapeutic Medicine

    Article Title: Oral immunization with rotavirus VP7-CTB fusion expressed in transgenic Arabidopsis thaliana induces antigen-specific IgA and IgG and passive protection in mice

    doi: 10.3892/etm.2018.6003

    Figure Lengend Snippet: Schematic of plant expression plasmids pPHAP1301-HRVVP7-CTB and pPHAP1301-HRVVP7. bar encodes a resistance marker for glufosinate selection. (A) HRVVP7 coding sequence. (Gly 4 Ser) 3 is the linker between HRVVP7 and CTB . (B) HRVVP7 coding sequence. HRVVP7, human rotavirus VP7; CTB, cholera toxin B subunit; Phas , β-phaseolin storage protein; 35s, Cauliflower mosaic virus; LB, left border; RB, right border; nos , nopaline synthase.

    Article Snippet: The intracellular rotavirus was detected using the mouse mAb directed against rotavirus VP7 (cat. no. C01715M; Meridian Life Science, Inc., Memphis, TN, USA; 1:500 dilution) as the primary antibody at 25°C for 30 min and fluorescein-isothiocyanate-labeled goat anti-mouse IgG antibody (cat. no. ab6785; Abcam) as the secondary antibody (1:500 dilution) at 37°C for 1 h. The cells were washed three times with PBS at room temperature (15 min/wash).

    Techniques: Expressing, CtB Assay, Marker, Selection, Sequencing

    HRVVP7-CTB molecular detection was performed using polymerase chain reaction and western blotting. Agarose gel electrophoresis was used to screen for positive transgenic lines following amplification. (A) HRVVP7-CTB fusion transgenic Arabidopsis thaliana lines. pUC19-HRVVP7-linker-CTB plasmid was used as the positive control (+). (B) HRVVP7 transgenic A. thaliana lines. pPHAP1301-HRVVP7 plasmid was used as the positive control (+). Proteins expressed by transgenic A. thaliana were assessed using western blotting. (C) HRVVP7-linker-CTB and (D) HRVVP7 expressed in seeds of T3 transgenic A. thaliana lines. HRVVP7, human rotavirus VP7; CTB, cholera toxin B subunit; WT, wild type A. thaliana; +, positive control; M, marker.

    Journal: Experimental and Therapeutic Medicine

    Article Title: Oral immunization with rotavirus VP7-CTB fusion expressed in transgenic Arabidopsis thaliana induces antigen-specific IgA and IgG and passive protection in mice

    doi: 10.3892/etm.2018.6003

    Figure Lengend Snippet: HRVVP7-CTB molecular detection was performed using polymerase chain reaction and western blotting. Agarose gel electrophoresis was used to screen for positive transgenic lines following amplification. (A) HRVVP7-CTB fusion transgenic Arabidopsis thaliana lines. pUC19-HRVVP7-linker-CTB plasmid was used as the positive control (+). (B) HRVVP7 transgenic A. thaliana lines. pPHAP1301-HRVVP7 plasmid was used as the positive control (+). Proteins expressed by transgenic A. thaliana were assessed using western blotting. (C) HRVVP7-linker-CTB and (D) HRVVP7 expressed in seeds of T3 transgenic A. thaliana lines. HRVVP7, human rotavirus VP7; CTB, cholera toxin B subunit; WT, wild type A. thaliana; +, positive control; M, marker.

    Article Snippet: The intracellular rotavirus was detected using the mouse mAb directed against rotavirus VP7 (cat. no. C01715M; Meridian Life Science, Inc., Memphis, TN, USA; 1:500 dilution) as the primary antibody at 25°C for 30 min and fluorescein-isothiocyanate-labeled goat anti-mouse IgG antibody (cat. no. ab6785; Abcam) as the secondary antibody (1:500 dilution) at 37°C for 1 h. The cells were washed three times with PBS at room temperature (15 min/wash).

    Techniques: CtB Assay, Polymerase Chain Reaction, Western Blot, Agarose Gel Electrophoresis, Transgenic Assay, Amplification, Plasmid Preparation, Positive Control, Marker

    Immunohistochemical analysis and viral challenge. Immunof-luorescent analysis of the virus-neutralizing effects of IgG antibodies directed against HRVVP7. (A) Negative control group, (B) HRVVP7-immunized group at 1:42 dilution, (C) HRVVP7-linker-CTB-immunized group at 1:42 dilution and (D) HRVVP7-linker-CTB-immunized group at 1:67 dilution. Magnification, ×125. (E) Incidence of diarrhea in neonatal suckling mice following challenge with simian rotavirus SA11. ***P

    Journal: Experimental and Therapeutic Medicine

    Article Title: Oral immunization with rotavirus VP7-CTB fusion expressed in transgenic Arabidopsis thaliana induces antigen-specific IgA and IgG and passive protection in mice

    doi: 10.3892/etm.2018.6003

    Figure Lengend Snippet: Immunohistochemical analysis and viral challenge. Immunof-luorescent analysis of the virus-neutralizing effects of IgG antibodies directed against HRVVP7. (A) Negative control group, (B) HRVVP7-immunized group at 1:42 dilution, (C) HRVVP7-linker-CTB-immunized group at 1:42 dilution and (D) HRVVP7-linker-CTB-immunized group at 1:67 dilution. Magnification, ×125. (E) Incidence of diarrhea in neonatal suckling mice following challenge with simian rotavirus SA11. ***P

    Article Snippet: The intracellular rotavirus was detected using the mouse mAb directed against rotavirus VP7 (cat. no. C01715M; Meridian Life Science, Inc., Memphis, TN, USA; 1:500 dilution) as the primary antibody at 25°C for 30 min and fluorescein-isothiocyanate-labeled goat anti-mouse IgG antibody (cat. no. ab6785; Abcam) as the secondary antibody (1:500 dilution) at 37°C for 1 h. The cells were washed three times with PBS at room temperature (15 min/wash).

    Techniques: Immunohistochemistry, Negative Control, CtB Assay, Mouse Assay

    Schematic of plant expression plasmids pPHAP1301-HRVVP7-CTB and pPHAP1301-HRVVP7. bar encodes a resistance marker for glufosinate selection. (A) HRVVP7 coding sequence. (Gly 4 Ser) 3 is the linker between HRVVP7 and CTB . (B) HRVVP7 coding sequence. HRVVP7, human rotavirus VP7; CTB, cholera toxin B subunit; Phas , β-phaseolin storage protein; 35s, Cauliflower mosaic virus; LB, left border; RB, right border; nos , nopaline synthase.

    Journal: Experimental and Therapeutic Medicine

    Article Title: Oral immunization with rotavirus VP7-CTB fusion expressed in transgenic Arabidopsis thaliana induces antigen-specific IgA and IgG and passive protection in mice

    doi: 10.3892/etm.2018.6003

    Figure Lengend Snippet: Schematic of plant expression plasmids pPHAP1301-HRVVP7-CTB and pPHAP1301-HRVVP7. bar encodes a resistance marker for glufosinate selection. (A) HRVVP7 coding sequence. (Gly 4 Ser) 3 is the linker between HRVVP7 and CTB . (B) HRVVP7 coding sequence. HRVVP7, human rotavirus VP7; CTB, cholera toxin B subunit; Phas , β-phaseolin storage protein; 35s, Cauliflower mosaic virus; LB, left border; RB, right border; nos , nopaline synthase.

    Article Snippet: The intracellular rotavirus was detected using the mouse mAb directed against rotavirus VP7 (cat. no. C01715M; Meridian Life Science, Inc., Memphis, TN, USA; 1:500 dilution) as the primary antibody at 25°C for 30 min and fluorescein-isothiocyanate-labeled goat anti-mouse IgG antibody (cat. no. ab6785; Abcam) as the secondary antibody (1:500 dilution) at 37°C for 1 h. The cells were washed three times with PBS at room temperature (15 min/wash).

    Techniques: Expressing, CtB Assay, Marker, Selection, Sequencing

    HRVVP7-CTB molecular detection was performed using polymerase chain reaction and western blotting. Agarose gel electrophoresis was used to screen for positive transgenic lines following amplification. (A) HRVVP7-CTB fusion transgenic Arabidopsis thaliana lines. pUC19-HRVVP7-linker-CTB plasmid was used as the positive control (+). (B) HRVVP7 transgenic A. thaliana lines. pPHAP1301-HRVVP7 plasmid was used as the positive control (+). Proteins expressed by transgenic A. thaliana were assessed using western blotting. (C) HRVVP7-linker-CTB and (D) HRVVP7 expressed in seeds of T3 transgenic A. thaliana lines. HRVVP7, human rotavirus VP7; CTB, cholera toxin B subunit; WT, wild type A. thaliana; +, positive control; M, marker.

    Journal: Experimental and Therapeutic Medicine

    Article Title: Oral immunization with rotavirus VP7-CTB fusion expressed in transgenic Arabidopsis thaliana induces antigen-specific IgA and IgG and passive protection in mice

    doi: 10.3892/etm.2018.6003

    Figure Lengend Snippet: HRVVP7-CTB molecular detection was performed using polymerase chain reaction and western blotting. Agarose gel electrophoresis was used to screen for positive transgenic lines following amplification. (A) HRVVP7-CTB fusion transgenic Arabidopsis thaliana lines. pUC19-HRVVP7-linker-CTB plasmid was used as the positive control (+). (B) HRVVP7 transgenic A. thaliana lines. pPHAP1301-HRVVP7 plasmid was used as the positive control (+). Proteins expressed by transgenic A. thaliana were assessed using western blotting. (C) HRVVP7-linker-CTB and (D) HRVVP7 expressed in seeds of T3 transgenic A. thaliana lines. HRVVP7, human rotavirus VP7; CTB, cholera toxin B subunit; WT, wild type A. thaliana; +, positive control; M, marker.

    Article Snippet: The intracellular rotavirus was detected using the mouse mAb directed against rotavirus VP7 (cat. no. C01715M; Meridian Life Science, Inc., Memphis, TN, USA; 1:500 dilution) as the primary antibody at 25°C for 30 min and fluorescein-isothiocyanate-labeled goat anti-mouse IgG antibody (cat. no. ab6785; Abcam) as the secondary antibody (1:500 dilution) at 37°C for 1 h. The cells were washed three times with PBS at room temperature (15 min/wash).

    Techniques: CtB Assay, Polymerase Chain Reaction, Western Blot, Agarose Gel Electrophoresis, Transgenic Assay, Amplification, Plasmid Preparation, Positive Control, Marker

    Expression of α‐amylase in normal human tissues. The mRNA expressions of α‐amylase in human normal tissues were quantified by real‐time PCR. The α‐amylase/GAPDH ratio for liver was set as the baseline value to which all transcript levels were normalized. mRNA of α‐amylase/GAPDH in human tissues I and II across Human MTC Panel I and II (A) and in human digestive tissues across the Human Digestive System MTC Panel (B). GAPDH, glyceraldehyde‐3‐phosphate dehydrogenase; mRNA, messenger RNA; MTC, multiple tissue cDNA; PCR, Polymerase chain reaction

    Journal: Journal of Cellular Biochemistry

    Article Title: α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation. α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation

    doi: 10.1002/jcb.29357

    Figure Lengend Snippet: Expression of α‐amylase in normal human tissues. The mRNA expressions of α‐amylase in human normal tissues were quantified by real‐time PCR. The α‐amylase/GAPDH ratio for liver was set as the baseline value to which all transcript levels were normalized. mRNA of α‐amylase/GAPDH in human tissues I and II across Human MTC Panel I and II (A) and in human digestive tissues across the Human Digestive System MTC Panel (B). GAPDH, glyceraldehyde‐3‐phosphate dehydrogenase; mRNA, messenger RNA; MTC, multiple tissue cDNA; PCR, Polymerase chain reaction

    Article Snippet: Thiazolyl blue tetrazolium bromide (MTT) was purchased from Sigma‐Aldrich Co. Rabbit anti‐α‐amylase immunoglobulin Gs (IgGs) to human pancreatic α‐amylase (anti‐HPA IgGs, K50894R) were purchased from Meridian Life Science, Inc (Memphis, TN).

    Techniques: Expressing, Real-time Polymerase Chain Reaction, Polymerase Chain Reaction

    Expression of α‐amylase in Caco‐2 cells cultured 28 days. The cells were seeded at 5 × 10 4 cells/cm 2 . The first medium change was made 2 days after seeding, and then the medium was changed every 2 to 3 days until 21 or 28 days. A, mRNA expression of α‐amylase: The cells were seeded and cultured in six‐well plates. The expression of α‐amylase was determined by real‐time PCR. The expression for days 3 to 28 (ΔΔ C t method vs that cultured for day 2) is shown; mean ± SE for 4 to 5 independent experiments. * P

    Journal: Journal of Cellular Biochemistry

    Article Title: α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation. α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation

    doi: 10.1002/jcb.29357

    Figure Lengend Snippet: Expression of α‐amylase in Caco‐2 cells cultured 28 days. The cells were seeded at 5 × 10 4 cells/cm 2 . The first medium change was made 2 days after seeding, and then the medium was changed every 2 to 3 days until 21 or 28 days. A, mRNA expression of α‐amylase: The cells were seeded and cultured in six‐well plates. The expression of α‐amylase was determined by real‐time PCR. The expression for days 3 to 28 (ΔΔ C t method vs that cultured for day 2) is shown; mean ± SE for 4 to 5 independent experiments. * P

    Article Snippet: Thiazolyl blue tetrazolium bromide (MTT) was purchased from Sigma‐Aldrich Co. Rabbit anti‐α‐amylase immunoglobulin Gs (IgGs) to human pancreatic α‐amylase (anti‐HPA IgGs, K50894R) were purchased from Meridian Life Science, Inc (Memphis, TN).

    Techniques: Expressing, Cell Culture, Real-time Polymerase Chain Reaction

    Identification of α‐amylase isozyme expressed in differentiated Caco‐2 cells. Caco‐2 cells were seeded at 5 × 10 4 cells/cm 2 and cultured in six‐well plates for 21 days. Total RNAs from cultured Caco‐2 cells were separately reacted with three sets of primers (#1, #2, #3) in RT‐PCR as described in Section 2. The RT‐PCR products were treated without (−) or with (+) Pst I, Hae II, or Bam HI. Arrowheads indicate the positions of migrated fragments that were cleaved (filled triangle) or not cleaved (open triangle), respectively. RT‐PCR, reverse transcription polymerase chain reaction

    Journal: Journal of Cellular Biochemistry

    Article Title: α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation. α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation

    doi: 10.1002/jcb.29357

    Figure Lengend Snippet: Identification of α‐amylase isozyme expressed in differentiated Caco‐2 cells. Caco‐2 cells were seeded at 5 × 10 4 cells/cm 2 and cultured in six‐well plates for 21 days. Total RNAs from cultured Caco‐2 cells were separately reacted with three sets of primers (#1, #2, #3) in RT‐PCR as described in Section 2. The RT‐PCR products were treated without (−) or with (+) Pst I, Hae II, or Bam HI. Arrowheads indicate the positions of migrated fragments that were cleaved (filled triangle) or not cleaved (open triangle), respectively. RT‐PCR, reverse transcription polymerase chain reaction

    Article Snippet: Thiazolyl blue tetrazolium bromide (MTT) was purchased from Sigma‐Aldrich Co. Rabbit anti‐α‐amylase immunoglobulin Gs (IgGs) to human pancreatic α‐amylase (anti‐HPA IgGs, K50894R) were purchased from Meridian Life Science, Inc (Memphis, TN).

    Techniques: Cell Culture, Reverse Transcription Polymerase Chain Reaction

    Immunostaining for α‐amylase in Caco‐2 cells. Caco‐2 cells were seeded at 5 × 10 4 cells/cm 2 and cultured on microcover glasses in 24‐well plates for 7 or 28 days. The cells were permeabilized after fixation and incubated with rabbit anti‐HPA IgGs (diluted to 1:150) and visualized by Alexa Flour488‐conjugated goat anti‐rabbit IgGs (diluted to 1:200, green fluorescence). Nuclei were counterstained with DAPI. Controls were incubated with only the secondary antibody without the anti‐HPA IgGs. The lower right magnification is ×6.67. Scale bars = 50 μm. DAPI, 4′,6‐diamidino‐2‐phenylindole; HPA, human pancreatic α‐amylase; IgG, immunoglobulin G

    Journal: Journal of Cellular Biochemistry

    Article Title: α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation. α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation

    doi: 10.1002/jcb.29357

    Figure Lengend Snippet: Immunostaining for α‐amylase in Caco‐2 cells. Caco‐2 cells were seeded at 5 × 10 4 cells/cm 2 and cultured on microcover glasses in 24‐well plates for 7 or 28 days. The cells were permeabilized after fixation and incubated with rabbit anti‐HPA IgGs (diluted to 1:150) and visualized by Alexa Flour488‐conjugated goat anti‐rabbit IgGs (diluted to 1:200, green fluorescence). Nuclei were counterstained with DAPI. Controls were incubated with only the secondary antibody without the anti‐HPA IgGs. The lower right magnification is ×6.67. Scale bars = 50 μm. DAPI, 4′,6‐diamidino‐2‐phenylindole; HPA, human pancreatic α‐amylase; IgG, immunoglobulin G

    Article Snippet: Thiazolyl blue tetrazolium bromide (MTT) was purchased from Sigma‐Aldrich Co. Rabbit anti‐α‐amylase immunoglobulin Gs (IgGs) to human pancreatic α‐amylase (anti‐HPA IgGs, K50894R) were purchased from Meridian Life Science, Inc (Memphis, TN).

    Techniques: Immunostaining, Cell Culture, Incubation, Fluorescence

    Effect of α‐amylase suppression on cell differentiation. Caco‐2 cells (0.5‐2.0 × 10 5 cells/cm 2 ) were transfected with none as a mock, nonspecific siRNA as a control, or the siRNA targeting α‐amylase. A, mRNA expression of α‐amylase. * P

    Journal: Journal of Cellular Biochemistry

    Article Title: α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation. α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation

    doi: 10.1002/jcb.29357

    Figure Lengend Snippet: Effect of α‐amylase suppression on cell differentiation. Caco‐2 cells (0.5‐2.0 × 10 5 cells/cm 2 ) were transfected with none as a mock, nonspecific siRNA as a control, or the siRNA targeting α‐amylase. A, mRNA expression of α‐amylase. * P

    Article Snippet: Thiazolyl blue tetrazolium bromide (MTT) was purchased from Sigma‐Aldrich Co. Rabbit anti‐α‐amylase immunoglobulin Gs (IgGs) to human pancreatic α‐amylase (anti‐HPA IgGs, K50894R) were purchased from Meridian Life Science, Inc (Memphis, TN).

    Techniques: Cell Differentiation, Transfection, Expressing

    Effect of α‐amylase suppression on cell proliferation. Caco‐2 cells (0.5‐2.0 × 10 5 cells/cm 2 ) were transfected with none as a mock, nonspecific siRNA as a control, or the siRNA targeting α‐amylase. A, Cell viability measured by MTT assay. The cell viability (% of mock) is shown as mean ± SE for three independent experiments. * P

    Journal: Journal of Cellular Biochemistry

    Article Title: α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation. α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation

    doi: 10.1002/jcb.29357

    Figure Lengend Snippet: Effect of α‐amylase suppression on cell proliferation. Caco‐2 cells (0.5‐2.0 × 10 5 cells/cm 2 ) were transfected with none as a mock, nonspecific siRNA as a control, or the siRNA targeting α‐amylase. A, Cell viability measured by MTT assay. The cell viability (% of mock) is shown as mean ± SE for three independent experiments. * P

    Article Snippet: Thiazolyl blue tetrazolium bromide (MTT) was purchased from Sigma‐Aldrich Co. Rabbit anti‐α‐amylase immunoglobulin Gs (IgGs) to human pancreatic α‐amylase (anti‐HPA IgGs, K50894R) were purchased from Meridian Life Science, Inc (Memphis, TN).

    Techniques: Transfection, MTT Assay

    Western‐blotting for α‐amylase and starch‐degrading activity in Caco‐2 cells and culture medium. Caco‐2 cells were seeded at 5 × 10 4 cells/cm 2 in dishes (150 × 20 mm) and cultured for 6 days. The cells and culture medium before and after culture (2 mL each) were prepared as described in Section 2. A, Western‐blotting for α‐amylase: The samples were boiled with SDS‐PAGE sample buffer including 2‐mercaptoethanol, and 10 μL aliquots were used for Western blot analysis as described in Section 2. Left, Protein staining using Sypro Ruby. Right, Immunostaining for α‐amylase using rabbit anti‐HPA IgGs and HRP‐conjugated goat anti‐rabbit IgGs, MW: molecular weight maker, Lane 1, purified pig pancreatic α‐amylase (0.8 μg/lane); 2, medium before culture (1.7 μg protein/lane); 3, medium after culture (1.8 μg protein/lane); 4, cell pellet (12 μg protein/lane); 5, cell extract (2.3 μg protein/lane). B, Starch‐degrading activity: Caco‐2 cells were cultured in dishes (150 × 20 mm) for 6 days. The cell pellet and extract were separately solubilized in 500 μL of 20 mM phosphate buffer, pH 6.9. The protein concentrations of the cell pellet and extract were 0.227 and 1.16 mg/mL, respectively. The cell pellet and extract (1, 10, and 100 times dilution) were used as enzyme samples (1 vol was 15 μL) as described in Section 2. HPA, human pancreatic α‐amylase; IgG, immunoglobulin G; SDS‐PAGE, sodium dodecyl sulfate‐polyacrylamide gel electrophoresis *** P

    Journal: Journal of Cellular Biochemistry

    Article Title: α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation. α‐Amylase expressed in human small intestinal epithelial cells is essential for cell proliferation and differentiation

    doi: 10.1002/jcb.29357

    Figure Lengend Snippet: Western‐blotting for α‐amylase and starch‐degrading activity in Caco‐2 cells and culture medium. Caco‐2 cells were seeded at 5 × 10 4 cells/cm 2 in dishes (150 × 20 mm) and cultured for 6 days. The cells and culture medium before and after culture (2 mL each) were prepared as described in Section 2. A, Western‐blotting for α‐amylase: The samples were boiled with SDS‐PAGE sample buffer including 2‐mercaptoethanol, and 10 μL aliquots were used for Western blot analysis as described in Section 2. Left, Protein staining using Sypro Ruby. Right, Immunostaining for α‐amylase using rabbit anti‐HPA IgGs and HRP‐conjugated goat anti‐rabbit IgGs, MW: molecular weight maker, Lane 1, purified pig pancreatic α‐amylase (0.8 μg/lane); 2, medium before culture (1.7 μg protein/lane); 3, medium after culture (1.8 μg protein/lane); 4, cell pellet (12 μg protein/lane); 5, cell extract (2.3 μg protein/lane). B, Starch‐degrading activity: Caco‐2 cells were cultured in dishes (150 × 20 mm) for 6 days. The cell pellet and extract were separately solubilized in 500 μL of 20 mM phosphate buffer, pH 6.9. The protein concentrations of the cell pellet and extract were 0.227 and 1.16 mg/mL, respectively. The cell pellet and extract (1, 10, and 100 times dilution) were used as enzyme samples (1 vol was 15 μL) as described in Section 2. HPA, human pancreatic α‐amylase; IgG, immunoglobulin G; SDS‐PAGE, sodium dodecyl sulfate‐polyacrylamide gel electrophoresis *** P

    Article Snippet: Thiazolyl blue tetrazolium bromide (MTT) was purchased from Sigma‐Aldrich Co. Rabbit anti‐α‐amylase immunoglobulin Gs (IgGs) to human pancreatic α‐amylase (anti‐HPA IgGs, K50894R) were purchased from Meridian Life Science, Inc (Memphis, TN).

    Techniques: Western Blot, Activity Assay, Cell Culture, SDS Page, Staining, Immunostaining, Molecular Weight, Purification, Polyacrylamide Gel Electrophoresis