- PA1-16987 - Invitrogen Antibodies DNM1L antibody

Submitted references CDDO-Me Attenuates Clasmatodendrosis in CA1 Astrocyte by Inhibiting HSP25-AKT Mediated DRP1-S637 Phosphorylation in Chronic Epilepsy Rats.

Lee DS, Kim TH, Park H, Kim JE
International journal of molecular sciences 2022 Apr 20;23(9)

β-Hydroxybutyrate Increases Exercise Capacity Associated with Changes in Mitochondrial Function in Skeletal Muscle.

Monsalves-Alvarez M, Morales PE, Castro-Sepulveda M, Sepulveda C, Rodriguez JM, Chiong M, Eisner V, Lavandero S, Troncoso R
Nutrients 2020 Jun 29;12(7)

CDDO-Me Selectively Attenuates CA1 Neuronal Death Induced by Status Epilepticus via Facilitating Mitochondrial Fission Independent of LONP1.

Kim JE, Park H, Choi SH, Kong MJ, Kang TC
Cells 2019 Aug 5;8(8)

PDI-mediated S-nitrosylation of DRP1 facilitates DRP1-S616 phosphorylation and mitochondrial fission in CA1 neurons.

Lee DS, Kim JE
Cell death & disease 2018 Aug 29;9(9):869

Mitochondrial Translocation of High Mobility Group Box 1 Facilitates LIM Kinase 2-Mediated Programmed Necrotic Neuronal Death.

Hyun HW, Ko AR, Kang TC
Frontiers in cellular neuroscience 2016;10:99

Endothelin-1 induces LIMK2-mediated programmed necrotic neuronal death independent of NOS activity.

Ko AR, Hyun HW, Min SJ, Kim JE, Kang TC
Molecular brain 2015 Oct 6;8:58

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of DNM1L using a polyclonal antibody (Product # PA1-16987).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of DNM1L in 0.5 mg/mL HeLa lysate. Samples were incubated in DNM1L polyclonal antibody (Product # PA1-16987). This experiment was performed under reducing conditions using the 12-230 kDa separation system.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of DNM1L in human HeLa and MCF7 cells, mouse 3T3 cells and rat PC12 cells. Samples were incubated in DNM1L polyclonal antibody (Product # PA1-16987) using a dilution of 2 µg/mL followed by an anti-rabbit HRP secondary antibody. Separated on a 7.5 % gel by SDS-PAGE, transferred to PVDF membrane and blocked in 5% non-fat milk in TBST. Detection: chemiluminescence.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis of DNM1L in DRP1 knockout, DRP1 wildtype MEFs. Samples were incubated in DNM1L polyclonal antibody (Product # PA1-16987 using a dilution of 1:500. Lane 1: DRP1 knockout; Lane 2: DRP1 wildtype MEFs.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of DNM1L in 0.5 mg/mL HeLa lysate. Samples were incubated in DNM1L polyclonal antibody (Product # PA1-16987). This experiment was performed under reducing conditions using the 12-230 kDa separation system.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockdown of DNM1L was achieved by transfecting HeLa with DNM1L specific siRNAs (Silencer® select Product # s19561, s19559). Western blot analysis (Fig. a) was performed using modified whole cell extracts (1% SDS) from the DNM1L knockdown cells (lane 3), non-specific scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blots were probed with DNM1L Polyclonal Antibody (Product # PA1-16987, 1:500 dilution) and Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25µg/mL, 1:4000 dilution). Densitometric analysis of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to DNM1L.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Western blot analysis was performed on modified whole cell extracts (1% SDS) (30 µg lysate) of SH-SY5Y (Lane 1), NTERA-2 (Lane 2), HeLa (Lane 3), HCT 116 (Lane 4), SK-OV-3 (Lane 5), NIH/3T3 (Lane 6), Raji (Lane 7), LNCaP (Lane 8), tissue extracts of Mouse Brain (Lane 9), Rat Brain (Lane 10) and Mouse Heart (Lane 11). The blot was probed with Anti-DNM1L Polyclonal Antibody (Product # PA1-16987, 1:500 dilution) and detected by chemiluminescence using Goat anti Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.25 µg/mL, 1:4000 dilution). A 82 kDa band corresponding to DNM1L was observed across cell lines and tissues tested.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunocytochemistry analysis of DNM1L in HeLa cells fixed for 10 minutes using 10% formalin and then permeabilized for 5 minutes using 1X TBS + 0.5% Triton X-100. Samples were incubated in DNM1L polyclonal antibody (Product # PA1-16987) using a dilution of 5 µg/mL overnight at 4 °C followed by anti-rabbit DyLight 488 (Green) with a dilution of 1:500. Alpha tubulin (DM1A) was used as a co-stain at 1:1000 and detected with an anti-mouse DyLight 550 (Red) at 1:500. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 40X objective.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunocytochemistry analysis of DNM1L in PC12 cells fixed for 10 minutes using 10% formalin and then permeabilized for 5 minutes using 1X PBS + 0.05% Triton-X100. Samples were incubated in DNM1L polyclonal antibody (Product # PA1-16987) using a dilution of 2 µg/mL overnight at 4 °C followed by anti-rabbit DyLight 488 (Green) with a dilution of 1:500. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 40X objective.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of DNM1L in renal tubular epithelium and visceral epithelial cells of the glomerulus. Samples were incubated in DNM1L polyclonal antibody (Product # PA1-16987). Human kidney cortex, 40X magnification.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Immunohistochemical analysis of DNM1L in formalin-fixed paraffin-embedded tissue section of mouse kidney. Samples were incubated in DNM1L polyclonal antibody (Product # PA1-16987) using a dilution of 1:300 followed by a HRP labeled secondary antibody and DAB reagent. Nuclei of the cells were counterstained with hematoxylin. This antibody generated a diffused cytoplasmic staining of DRP1 in the epithelial cells of various tubules and in the cells of glomeruli.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Flow cytometry of DNM1L in HeLa cells (blue) and a matched isotype control (orange). Samples were incubated in DNM1L polyclonal antibody (Product # PA1-16987) using a dilution of 5 µg/mL for 30 minutes at room temperature. Cells were fixed with 4% PFA and then permeabilized with 0.1% saponin. Both antibodies were conjugated to Alexa Fluor 488.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry of DNM1L in HeLa cells and a matched isotype control. Samples were incubated in DNM1L polyclonal antibody (Product # PA1-16987) using a dilution of 2.5 µg/mL for 30 minutes at room temperature followed by a Rabbit IgG (H+L) Cross-Adsorbed Secondary Antibody, Dylight™ 550. Cells were fixed with 4% PFA and then permeabilized with 0.1% saponin.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 1 The effects of l -NAME on SNO-PDI, SNO-DRP1, and mitochondrial fusion in CA1 neurons under normal condition. l -NAME inhibits NO generation and the binding of PDI to DRP1, and reduces SNO-PDI and SNO-DRP1 levels without changed PDI and DRP1 expressions. However, l -NAME elongates mitochondrial length in CA1 neurons, accompanied by elevated DRP1-S616 phosphorylation. a Quantitative values (mean +- S.E.M) of nitrate/nitrite levels in the hippocampus 7 days after l -NAME infusion. Open circles indicate each individual value. Horizontal bars indicate mean value. Error bars indicate SEM ( *p < 0.05 vs. non-SE animals; n = 7, respectively). b Representative western blot for co-immunoprecipitation of PDI interaction with DRP1 following l -NAME treatment. c Quantification of co-immunoprecipitation analyses of PDI interactions with DRP1 following l -NAME treatment. Open circles indicate each individual value. Horizontal bars indicate mean value. Error bars indicate SEM ( *p < 0.05 vs. vehicle; n = 7, respectively). d Representative western blot for expression, S -nitrosylation and phosphorylation levels of PDI and DRP1 following l -NAME treatment. e - g Quantification of values (mean +- S.E.M) of expression, phosphorylation, and S -nitrosylation levels of PDI and DRP1 following l -NAME treatment. Open circles indicate each individual value. Horizontal bars indicate mean value. Error bars indicate SEM ( *p < 0.05 vs. vehicle; n = 7, respectively). h

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 4 The effects of l -NAME on SNO-PDI, SNO-DRP1, Trx1 activity, and mitochondrial fusion in CA1 neurons 3 days after SE. SE significantly reduces expression levels of DRP1 and PDI, PDI-binding to DRP1, and phosphorylation levels of DRP1-S616 and S637 levels independent of Trx1 activity. Both SNO-PDI and SNO-DRP1 are more diminished by l -NAME treatment. a Representative western blot for the effect of l -NAME on expression, S -nitrosylation, and phosphorylation levels of PDI and DRP1 following SE. b - d Quantification of values (mean +- S.E.M) of expression, phosphorylation, and S -nitrosylation levels of PDI and DRP1. Open circles indicate each individual value. Horizontal bars indicate mean value. Error bars indicate SEM (*, # p < 0.05 vs. non-SE animals and vehicle, respectively; n = 7, respectively). e Quantification of values (mean +- S.E.M) of Trx1 activity. Open circles indicate each individual value. Horizontal bars indicate mean value. Error bars indicate SEM ( n = 7, respectively). f Representative western blot for the effect of l -NAME on co-immunoprecipitation of PDI interaction with DRP1 following SE. g Quantification of co-immunoprecipitation analyses of PDI interactions with DRP1. Open circles indicate each individual value. Horizontal bars indicate mean value. Error bars indicate SEM ( *p < 0.05 vs. non-SE animals; n = 7, respectively)

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 6 The effects of PDI knockdown on SNO-PDI, SNO-DRP1, and mitochondrial fusion in CA1 neurons under normal condition. PDI knockdown reduces PDI expression, the binding of PDI to DRP1, SNO-PDI, and SNO-DRP1 levels without changed DRP1 expression and nitrate/nitrite concentration. However, PDI knockdown elongates mitochondrial length in CA1 neurons, accompanied by elevated DRP1-S616 phosphorylation. a Quantitative values (mean +- S.E.M) of nitrate/nitrite levels in the hippocampus 7 days after PDI siRNA infusion. Open circles indicate each individual value. Horizontal bars indicate mean value. Error bars indicate SEM ( n = 7, respectively). b Representative western blot for co-immunoprecipitation of PDI interaction with DRP1 following PDI knockdown. c Quantification of co-immunoprecipitation analyses of PDI interactions with DRP1 following PDI knockdown. Open circles indicate each individual value. Horizontal bars indicate mean value. Error bars indicate SEM ( *p < 0.05 vs. control siRNA; n = 7, respectively). d Representative western blot for expression, S -nitrosylation, and phosphorylation levels of PDI and DRP1 following PDI knockdown. e - g Quantification of values (mean +- S.E.M) of expression, phosphorylation, and S -nitrosylation levels of PDI and DRP1 following PDI knockdown. Open circles indicate each individual value. Horizontal bars indicate mean value. Error bars indicate SEM ( *p < 0.05 vs. control siRNA; n = 7, respectively). h Quantifica

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig. 6 Effect of BQ788 and Y-27632 on dysfunction of mitochondrial fission at 3 days after SE. a Western blot images of DRP1, DRP1 S616 and DRP1 S637 in the hippocampus. As compared to vehicle, both BQ788 and Y-27632 (a ROCK inhibitor) attenuate the reductions in DRP1 and DRP1 S616 expression, but increase DRP1 S637 expression. b Quantitative values (mean +- S.E.M) of DRP1, DRP1 S616, DRP1 S637 level ( n = 10 per each group). Significant differences from vehicle, * p < 0.05. c Quantitative values (mean +- S.E.M) of DRP1 S616/S637 ratio in the hippocampus ( n = 10 per each group). Significant differences from vehicle, * p < 0.05. d Quantitative values (mean +- S.E.M) of mitochondrial length in the CA1 neurons ( n = 10 per each group). Significant differences from non-SE animals, * p < 0.05 e-h Representative photographs of mitochondria and NeuN in the CA1 neurons. SE increases mitochondrial length and sphere formation. Both BQ788 and Y-27632 alleviate mitochondrial elongation and sphere formation induced by SE. Bar = 6.25 mum

Supportive validation

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Experimental details: Figure 4 Effect of LMIK2 siRNA and LMB on HMGB1 expression, DRP1 expression and DRP phosphorylation level at 3 days after SE. (A) Western blot images of HMGB1, DRP1, DRP1 S616 and DRP1 S637 in the hippocampus. Both LIMK2 siRNA and LMB abolish the reduction of HMGB1 release induced by SE. Only LIMK2 siRNA prevents reductions in DRP1 expression, DRP1 S616 level and DRP1 S616/S637 level induced by SE. (B) Quantitative values (mean +- SEM) of HMGB1, DRP1, DRP1 S616, DRP1 S637 level, based on western blot ( n = 7, respectively). * p < 0.05 vs. control siRNA and vehicle, respectively.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2 Six weeks of beta-HB intake alter the electron transport chain and mitochondrial dynamics protein levels. ( A ) Representative OXPHOS Western blot from Tibialis anterior whole tissue extract. ( B ) Tibialis Anterior OXPHOS quantification. ( C ) mHsp70 levels. ( D ) Tibialis anterior mitochondrial fission and fusion proteins representative Western blot. ( E ) Mfn2. ( F ) Opa1 and ( G ) Drp1. ( H ) Soleus representative OXPHOS Western blot. ( I ) Soleus OXPHOS quantification. ( J ) mHsp70. ( K ) Soleus mitochondrial fission and fusion proteins representative Western blot. ( L ) Mfn2. ( M ) Opa1 and ( N ) Drp1. The grouping of blots cropped from different parts of the same gel were divided by black lines. n = 5-6 mice per condition, * p < 0.05, Unpaired t -Test. Values expressed as mean +- SEM.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Effects of CDDO-Me and 3CAI on HSP25, AKT S473, DRP1 S616 and DRP1 S637 levels in the stratum radiatum of the CA1 region of epileptic rats. As compared to vehicle (Veh), CDDO-Me (CDDO) reduces HSP25, AKT S473 and DRP1 S637 levels. 3CAI decreases AKT S473 and DRP1 S637 levels. ( A ) Representative Western blot images of HSP25, AKT, AKT S473, DRP1, DRP1 S616 and DRP1 S637. ( B - E ) Quantifications of HSP25 ( B ), AKT S473 ( C ), DRP1 S616 ( D ) and DRP1 S637 levels. Open circles indicate each value. Horizontal bars indicate the mean value. Error bars indicate SEM (* p < 0.05 vs. Vehicle, respectively; n = 7, respectively).

PA1-16987

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