Invitrogen

PDLIM5 Polyclonal Antibody

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PDLIM5 Antibody in Immunocytochemistry (ICC/IF) — Immunofluorescence analysis of ENH was performed using 70% confluent log phase MCF-7 cells. The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, and blocked with 1% BSA for 1 hour at room temperature. The cells were labeled with ENH Rabbit Polyclonal Antibody (Product # 38-8800) at 2 µg/mL in 0.1% BSA and incubated for 3 hours at room temperature and then labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conj µgate (Product # A27034) at a dilution of 1:2000 for 45 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant with DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor® 555 Rhodamine Phalloidin (Product # R415, 1:300). Panel d represents the merged image showing cytoplasmic and cell junctional localization. Panel e shows the no primary antibody control. The images were captured at 60X magnification.

PDLIM5 Antibody in Western Blot (WB) — Western blot analysis was performed on whole cell extracts of C2C12 (Lane 1), PC-3 (Lane 2), Caco-2 (Lane 3) and U-87 MG (Lane 4). The blots were probed with Anti- ENH Rabbit Polyclonal Antibody (Product # 38-8800, 2 µg/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, HRP conj µgate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). Bands of ~ 64 and 54 kDa corresponding to ENH were observed across cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody using iBind™ Flex Western Starter Kit (Product # SLF2000S). Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

PDLIM5 Antibody in Western Blot (WB) — Western blot analysis of (A) rat heart homogenates, (B) PC-3, (C) LNCap-FGC, (D) Caco-2, (E) HT-29 cell lysates using Rb anti-ENH (Product # 38-8800)

PDLIM5 Antibody in Immunocytochemistry (ICC/IF) — Figure 6. Delta-catenin interacts with Magi1 and Pdlim5 in developing hippocampal neurons. (A) Expression of endogenous delta-catenin (green), Magi1 (red, top), and Pdlim5 (red, bottom) in developing primary rat hippocampal neurons at 1 DIV, 7 DIV, and 28 DIV. Scale bars, 20 um. (B and C) Detection of direct associations between endogenous delta-catenin and the proteins Magi1 and Pdlim5 in developing hippocampal neurons. A PLA (Duolink), in combination with delta-catenin, Magi1, and Pdlim5 antibodies, was used to detect direct endogenous protein-protein interactions in 7-DIV hippocampal neurons. (B) Visualization of Magi1:delta-catenin (left) and Pdlim5:delta-catenin (center) interactions, as detected by the presence of red puncta. Insets show presence of puncta on/near small protrusions from the cell in several cases. Antibodies specific to delta-catenin and c-Jun were used in a negative control assay (right). Scale bars, 20 um in main panels and 5 um in zoomed insets. (C) Quantification of average puncta per neuron in each condition (delta-catenin+ Magi1: 16.01 [P = 0.0159]; Pdlim5: 37.73 [P = 0.0007]; c-Jun/Control: 2.393); n = 3 independent experiments. (D) Quantification of puncta/cell detected by a PLA (Duolink) in 7-DIV rat hippocampal neurons treated with DHPG. Cells were fixed immediately following treatment (no treatment versus 20 uM DHPG; 10 min) and subjected to the assay. DHPG treatment produced an increase in Pdlim5:delta-catenin puncta relative to no treatment

PDLIM5 Antibody in Western Blot (WB) — Figure S3. shRNA-mediated KD of delta-catenin, Magi1, and Pdlim5. (A) Immunoblots of delta-catenin, Magi1, and Pdlim5 from hippocampal neuron culture cell lysates from 1 DIV to 28 DIV. (B) Immunoblots of cell lysates from HEK293 cells coexpressing HA-delta-catenin, FLAG-Magi1, or Myc-Pdlim5 with either a control vector or the appropriate shRNA. We find that delta-catenin shRNA results in a near-complete loss of delta-catenin (~100% KD), while the Magi1 (~68% KD) and Pdlim5 (~60% KD) shRNAs produce slightly less complete KDs of their respective proteins. (C-E) Analysis of neurons expressing Magi1 shRNA compared with rescues (neurons expressing shRNA-KD and cDNA-overexpression constructs). (C) Expression of Flag-tagged Magi1 in Magi1 shRNA-expressing neurons resulted in a rescue of the shRNA dendrite length phenotype (control: 35.96 +- 1.56 um; shRNA: 21.72 +- 0.98 um; rescue: 33.94 +- 1.44 um, P < 0.0001). (D) Expression of Magi1 cDNA in combination with Magi1 shRNA did not produce a significant impact on dendrite density (control: 16.69 +- 1.26; shRNA: 17.92 +- 2.01; rescue: 18.22 +- 0.85 um, P = 0.7911). (E) Sholl analysis of Magi1 shRNA and rescue neurons reveals that rescue of the shRNA via expression of Magi1 results in a dendritic arborization pattern similar to that of controls. (F-H) Analysis of neurons expressing Pdlim5 shRNA compared with rescues (neurons expressing shRNA-KD and cDNA-overexpression constructs). (F) Expression of Myc-tagged Pdlim5 in Pdlim5 shRNA-expr

Product Details

38-8800

Applications	Tested Dilution	Publications
Western Blot (WB)	2 µg/mL	View 1 publication 1 publication
Immunocytochemistry (ICC/IF)	2 µg/mL	View 1 publication 1 publication
in situ PLA (PLA)	-	View 1 publication 1 publication

Product Specifications
Species Reactivity	Rat, Human, Mouse
Published species	Human
Host/Isotype	Rabbit / IgG
Class	Polyclonal
Type	Antibody
Immunogen	Synthetic peptide derived from the N-terminal region of the human ENH (enigma homologue 1, LIM protein, protein kinase C- binding protein enigma) protein, which differs from mouse and rat ENH by one amino acid. View immunogen
Conjugate	Unconjugated
Form	Liquid
Concentration	0.25 mg/mL
Purification	Antigen affinity chromatography
Storage buffer	PBS, pH 7.4
Contains	0.1% sodium azide
Storage conditions	-20°C
RRID	AB_2533388

Target Information

May play an important role in the heart development by scaffolding PKC to the Z-disk region. May play a role in the regulation of cardiomyocyte expansion. Overexpression promotes the development of heart hypertrophy. Contributes to the regulation of dendritic spine morphogenesis in neurons. May restrain postsynaptic growth of excitatory synapses.

For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.

Product References

Novel phospho-switch function of delta-catenin in dendrite development.

The Journal of cell biology

Baumert R,Ji H,Paulucci-Holthauzen A,Wolfe A,Sagum C,Hodgson L,Arikkath J,Chen X,Bedford MT,Waxham MN,McCrea PD

38-8800 was used in Proximity Ligation Assay (PLA), Immunocytochemistry to report a new mechanism instructing dendrites to branch versus extend.

Mon Nov 02 00:00:00 EST 2020

Novel phospho-switch function of delta-catenin in dendrite development.

The Journal of cell biology

Baumert R,Ji H,Paulucci-Holthauzen A,Wolfe A,Sagum C,Hodgson L,Arikkath J,Chen X,Bedford MT,Waxham MN,McCrea PD

38-8800 was used in Proximity Ligation Assay (PLA), Immunocytochemistry to report a new mechanism instructing dendrites to branch versus extend.

Mon Nov 02 00:00:00 EST 2020

Novel phospho-switch function of delta-catenin in dendrite development.

The Journal of cell biology

Baumert R,Ji H,Paulucci-Holthauzen A,Wolfe A,Sagum C,Hodgson L,Arikkath J,Chen X,Bedford MT,Waxham MN,McCrea PD

38-8800 was used in Proximity Ligation Assay (PLA), Immunocytochemistry to report a new mechanism instructing dendrites to branch versus extend.

Mon Nov 02 00:00:00 EST 2020

Novel phospho-switch function of delta-catenin in dendrite development.

The Journal of cell biology

Baumert R,Ji H,Paulucci-Holthauzen A,Wolfe A,Sagum C,Hodgson L,Arikkath J,Chen X,Bedford MT,Waxham MN,McCrea PD

38-8800 was used in Proximity Ligation Assay (PLA), Immunocytochemistry to report a new mechanism instructing dendrites to branch versus extend.

Mon Nov 02 00:00:00 EST 2020

Novel phospho-switch function of delta-catenin in dendrite development.

The Journal of cell biology

Baumert R,Ji H,Paulucci-Holthauzen A,Wolfe A,Sagum C,Hodgson L,Arikkath J,Chen X,Bedford MT,Waxham MN,McCrea PD

38-8800 was used in Proximity Ligation Assay (PLA), Immunocytochemistry to report a new mechanism instructing dendrites to branch versus extend.

Mon Nov 02 00:00:00 EST 2020

Bioinformatics

Protein Aliases: Enigma homolog; enigma-like LIM domain protein; Enigma-like PDZ and LIM domains protein; PDZ and LIM domain protein 5

Gene Aliases: 1110001A05Rik; AI987914; C87059; ENH; ENH1; Enh2; Enh3; L9; LIM; PDLIM5

UniProt ID: (Human) Q96HC4, (Mouse) Q8CI51, (Rat) Q62920

Entrez Gene ID: (Human) 10611, (Mouse) 56376, (Rat) 64353

Molecular Function: actin or actin-binding cytoskeletal protein

regulation of synapse assembly regulation of dendritic spine morphogenesis cell growth involved in cardiac muscle cell development cell-cell adhesion heart development

Process(es)

actin binding protein kinase C binding protein binding zinc ion binding actinin binding protein N-terminus binding cadherin binding involved in cell-cell adhesion metal ion binding receptor signaling complex scaffold activity