- 45-6600 - Invitrogen Antibodies PDHA1 antibody

Submitted references Constitutive activation of the PI3K-Akt-mTORC1 pathway sustains the m.3243 A > G mtDNA mutation.

Chung CY, Singh K, Kotiadis VN, Valdebenito GE, Ahn JH, Topley E, Tan J, Andrews WD, Bilanges B, Pitceathly RDS, Szabadkai G, Yuneva M, Duchen MR
Nature communications 2021 Nov 4;12(1):6409

Antioxidant-Conjugated Peptide Attenuated Metabolic Reprogramming in Pulmonary Hypertension.

Varghese MV, Niihori M, Eccles CA, Kurdyukov S, James J, Rafikova O, Rafikov R
Antioxidants (Basel, Switzerland) 2020 Jan 25;9(2)

Inhibition of Anaplerosis Attenuated Vascular Proliferation in Pulmonary Arterial Hypertension.

Valuparampil Varghese M, James J, Eccles CA, Niihori M, Rafikova O, Rafikov R
Journal of clinical medicine 2020 Feb 6;9(2)

Synaptic Activity Drives a Genomic Program That Promotes a Neuronal Warburg Effect.

Bas-Orth C, Tan YW, Lau D, Bading H
The Journal of biological chemistry 2017 Mar 31;292(13):5183-5194

Modulation of astrocytic mitochondrial function by dichloroacetate improves survival and motor performance in inherited amyotrophic lateral sclerosis.

Miquel E, Cassina A, Martínez-Palma L, Bolatto C, Trías E, Gandelman M, Radi R, Barbeito L, Cassina P
PloS one 2012;7(4):e34776

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis was performed on whole cell extracts (30 µg lysate) of Hep G2 (Lane 1), A-431 (Lane 2), U-87 MG (Lane 3), MOLT-4 (Lane 4), NIH/3T3 (Lane 5) and Neuro-2a (Lane 6). The blot was probed with Anti-PDH Monoclonal Antibody (Product # 45-6600, 1 µg/mL) and detected by chemiluminescence using Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.25 µg/mL, 1:4000 dilution). A 40 kDa band corresponding to PDH was observed across the cell lines tested.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of PDH in heart mitochondria extracts using a PDH Monoclonal antibody (Product # 45-6600) at a concentration of 1 µg/mL. Lane 1: Isolated mitochondria from human heart at 5 µg, Lane 2: Isolated mitochondria from bovine heart at 1 µg, Lane 3: Isolated mitochondria from rat heart at 10 µg, Lane 4: Isolated mitochondria from mouse heart at 10 µg, Lane 5: HepG2 cell lysate at 20 µg. Predicted band size: 43 kDa.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunofluorescent analysis of PDH in HeLa cells using a PDH Monoclonal antibody (Product # 45-6600) at 1 µg/mL. Samples were fixed with 4% paraformaldehyde and permeabilized with Triton X-100. The secondary antibody was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 dilution, seen in green. DAPI was used to stain the cell nuclei as seen in blue.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of PDH in HeLa cells using a PDH Monoclonal antibody (Product # 45-6600) at 1 µg/mL. Samples were fixed with 4% paraformaldehyde and permeabilized with Triton X-100. The secondary antibody was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 dilution, seen in green. DAPI was used to stain the cell nuclei as seen in blue.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometric analysis of PDH in HL-60 cells using a PDH Monoclonal antibody (Product # 45-6600) at 1 µg/mL, as seen in blue. Isotype control antibody as seen in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2 DCA recovers mitochondrial respiration rate and controls proliferation in SOD1 G93A astrocytes. A) Representative immunoblot for PDH-E1alpha(pSer 293 ), total PDH-E1alpha, and beta-actin of lysates from non Tg and SOD1 G93A astrocytes after 24 h treatment with DCA or vehicle as described in Methods . B) Quantification of the PDH-E1alpha(pSer 293 ) to total PDH-E1alpha ratio between relative densitometric levels normalized against vehicle-treated non Tg astrocytes. C) Calculated respiratory control ratio (RCR) for mitochondria from non Tg or SOD1 G93A -bearing astrocytes treated with DCA or vehicle as indicated. D) Percentage of BrdU immunoreactive nuclei of non Tg and SOD1 G93A astrocytes after 24 h treatment with DCA. Data for panels B, C, and D are expressed as mean +- SEM from three independent experiments performed in duplicate. *p

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig. 1 The m.3243 A > G mtDNA mutation causes mitochondrial dysfunction and oxidative stress. a PCR-RFLP and ARMS-qPCR were used to quantify the mutation load for patient-derived fibroblasts ( n = 3 independent biological samples) and A549 cybrid cells ( n = 5 independent biological samples). b - d Cell respiratory capacity was measured using the Seahorse XFe96 extracellular flux analyser in patient fibroblasts ( n = 6 culture wells) showing a major decrease in oxygen consumption under all conditions ( b , p < 0.0001). Oxygen consumption dependent on ATP production ( c , p < 0.0001)--the response to oligomycin--and extracellular acidification rate (ECAR) are plotted ( d , p < 0.0001). e , f The expression of respiratory chain proteins and supercomplex assembly in patient fibroblasts were assessed using blue native gel electrophoresis (BNGE, e ) and quantified ( f , p < 0.0001), showing a major decrease in the assembly of all supercomplexes from the patient-derived cells ( n = 3 independent experiments). g , h The mitochondrial membrane potential of fibroblasts was measured using TMRM with confocal microscopy ( g ) and quantified ( h , p < 0.0001), showing a significant decrease in potential in patient-derived cells ( n = 5 independent experiments). Scale bar = 50 mum. i ROS production rates of control and patient fibroblasts were measured using the reporters, DHE ( p < 0.0001) and MitoSOX ( p < 0.0001). Rates of ROS production were significantly increased in patient-derived c

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 5 NP treatment balanced glucose influx and glycolysis in PAH. (A ) Western blot of total glucose transporter Glut4 expression in control, SU2, and NP treatment. ( B ) The ratio of Glut4 expression in membrane/cytosol was found significantly increased in SU2, and NP treatment controlled Glut4 translocation. ( C,D ) HK1 and GAPDH expression increased in sugen/hypoxia treatment represents increased glucose utilization by glycolysis. ( E ) Lactate dehydrogenase expression elevated in SU2 represents increased lactate production in early PAH. ( F ) Increased lactate dehydrogenase activity in lung tissue was attenuated with NP treatment. ( G,H ) Pyruvate dehydrogenase expression and activity were found significantly increased in NP-treated sugen/hypoxia. Data expressed as mean +- SE normalized on the total protein, N = 6-8, * p < 0.05 versus control, # p < 0.05 versus SU2 by ANOVA.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 5 PC inhibitor controlled the balance between lactate dehydrogenase (LDH) and pyruvate dehydrogenase (PDH) in PAH. ( A ) Lactate dehydrogenase (LDHA) increased in the SU5 group was significantly reduced with PC inhibitor treatment. ( B ) Total PDH expression was not significantly altered, but ( C ) PDH activity significantly attenuated in the SU5 PAH group, and PC inhibitor treatment reduced this back to the control level. Data expressed as box-and-whisker plot with median value +- min max, * p < 0.05 versus control, # p < 0.05 versus SU5 by ANOVA.

45-6600

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