Phospho-PAK6 (Ser560) Polyclonal Antibody

$Paraformaldehyde-fixed, paraffin embedded (Rat skin); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with (PAK4(Ser474)\/PAK5(Ser602)\/PAK6(Ser560)) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:200 overnight at 4\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.$

Paraformaldehyde-fixed, paraffin embedded (Rat skin); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with (PAK4(Ser474)\/PAK5(Ser602)\/PAK6(Ser560)) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:200 overnight at 4\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

$Paraformaldehyde-fixed, paraffin embedded (Rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with (PAK4(Ser474)\/PAK5(Ser602)\/PAK6(Ser560)) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:200 overnight at 4\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.$

Paraformaldehyde-fixed, paraffin embedded (Rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with (PAK4(Ser474)\/PAK5(Ser602)\/PAK6(Ser560)) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:200 overnight at 4\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

$Paraformaldehyde-fixed, paraffin embedded (mouse brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with (PAK4(Ser474)\/PAK5(Ser602)\/PAK6(Ser560)) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:200 overnight at 4\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.$

Paraformaldehyde-fixed, paraffin embedded (mouse brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with (PAK4(Ser474)\/PAK5(Ser602)\/PAK6(Ser560)) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:200 overnight at 4\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

$Paraformaldehyde-fixed, paraffin embedded (mouse skin); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with (PAK4(Ser474)\/PAK5(Ser602)\/PAK6(Ser560)) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:100 overnight at 4\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.$

Paraformaldehyde-fixed, paraffin embedded (mouse skin); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with (PAK4(Ser474)\/PAK5(Ser602)\/PAK6(Ser560)) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:100 overnight at 4\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

$Paraformaldehyde-fixed, paraffin embedded (mouse esophageal); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with (PAK4(Ser474)\/PAK5(Ser602)\/PAK6(Ser560)) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:100 overnight at 4\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.$

Paraformaldehyde-fixed, paraffin embedded (mouse esophageal); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37\u00b0C for 30min; Antibody incubation with (PAK4(Ser474)\/PAK5(Ser602)\/PAK6(Ser560)) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:100 overnight at 4\u00b0C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.

$Lane 1: Human PANC-1 cell lysates; Lane 2: Human A431 cell lysates; Lane 3: Human A549 cell lysates; Lane 4: Human MCF-7 cell lysates probed with Phospho-PAK6 (Ser560) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:1000 dilution and 4\u00b0C overnight incubation. Followed by conjugated secondary antibody incubation at 1:20000 for 60 min at 37˚C.$

Lane 1: Human PANC-1 cell lysates; Lane 2: Human A431 cell lysates; Lane 3: Human A549 cell lysates; Lane 4: Human MCF-7 cell lysates probed with Phospho-PAK6 (Ser560) Polyclonal Antibody, Unconjugated (bs-3314R) at 1:1000 dilution and 4\u00b0C overnight incubation. Followed by conjugated secondary antibody incubation at 1:20000 for 60 min at 37˚C.

$A-431 cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum, C-0005) at 37\u00b0C for 20 min; Antibody incubation with (Phospho-PAK6 (Ser560)) polyclonal Antibody, Unconjugated (bs-3314R) 1:100, 90 minutes at 37\u00b0C; followed by a conjugated Goat Anti-Rabbit IgG antibody at 37\u00b0C for 90 minutes, DAPI (blue, C02-04002) was used to stain the cell nuclei.$

A-431 cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum, C-0005) at 37\u00b0C for 20 min; Antibody incubation with (Phospho-PAK6 (Ser560)) polyclonal Antibody, Unconjugated (bs-3314R) 1:100, 90 minutes at 37\u00b0C; followed by a conjugated Goat Anti-Rabbit IgG antibody at 37\u00b0C for 90 minutes, DAPI (blue, C02-04002) was used to stain the cell nuclei.

$A594 cells were\u00a0fixed with 4% PFA for 10min at room temperature,permeabilized with\u00a090% ice-cold methanol for 20 min at -20\u2103,\u00a0and incubated in 5% BSA blocking buffer\u00a0for 30 min at room temperature. Cells were then stained with Phospho-PAK6 (Ser560) Antibody(bs-3314R)at 1:100 dilution\u00a0in blocking buffer and\u00a0incubated for 30 min at\u00a0room temperature,\u00a0washed twice with 2%BSA in PBS,\u00a0followed by\u00a0secondary antibody incubation\u00a0for 40 min\u00a0at\u00a0room temperature. Acquisitions of 20,000 events were performed.\u00a0Cells stained with primary antibody\u00a0(green), and isotype control (orange).$

A594 cells were\u00a0fixed with 4% PFA for 10min at room temperature,permeabilized with\u00a090% ice-cold methanol for 20 min at -20\u2103,\u00a0and incubated in 5% BSA blocking buffer\u00a0for 30 min at room temperature. Cells were then stained with Phospho-PAK6 (Ser560) Antibody(bs-3314R)at 1:100 dilution\u00a0in blocking buffer and\u00a0incubated for 30 min at\u00a0room temperature,\u00a0washed twice with 2%BSA in PBS,\u00a0followed by\u00a0secondary antibody incubation\u00a0for 40 min\u00a0at\u00a0room temperature. Acquisitions of 20,000 events were performed.\u00a0Cells stained with primary antibody\u00a0(green), and isotype control (orange).

Applications

WB
IHC-P
IHC-F
IF
Flow-Cyt
ICC/IF

Reactivity

Human
Mouse
Rat

Overview
Catalog #	bs-3314R
Product Name	Phospho-PAK6 (Ser560) Polyclonal Antibody
Applications	WB, IHC-P, IHC-F, IF, Flow-Cyt, ICC/IF
Reactivity	Human, Mouse, Rat
Specifications
Conjugation	Unconjugated
Host	Rabbit
Source	KLH conjugated synthetic phosphopeptide derived from human PAK4 around the phosphorylation site of PAK4 Ser474 [RK(p-S)LV]
Modification Site	Ser560
Clonality	Polyclonal
Clone #	#REF!
Isotype	IgG
Concentration	1ug/ul
Purification	Purified by Protein A.
Storage Buffer	0.01M TBS(pH7.4) with 1% BSA, 0.02% Proclin300 and 50% Glycerol.
Storage Condition	Store at -20C for 12 months.
Target
Gene ID	10298
Synonyms	PAK4Ser474; PAK5Ser602; PAK6Ser560; p21 activated kinase 4 ; p21CDKN1A activated kinase 4; PAK 4; Protein kinase related to S.cerevisiae STE20 effector for Cdc42Hs; Serine threonine kinase PAK 4; Serine/threonine protein kinase PAK 4; Serine/threonine protein kinase PAK4; KIAA1142; PAK4_HUMAN.
Background	p21-activated kinases (PAKs) belong to the family of serine/threonine kinases involved in the control of various cellular processes, including the cell cycle, dynamics of the cytoskeleton, apoptosis, oncogenic transformation, and transcription. All PAK family members are characterized by the presence of p21-binding domain. p21-activated kinases are regulated by the small GTP-binding proteins Rac and Cdc42, and lipids, which stimulate autophosphorylation and phosphorylation of exogenous substrates. Serine (Ser-474) is the likely autophosphorylation site in the kinase domain of PAK4 in vivo. Phosphospecific directed against serine 474 detect activated PAK4 on the Golgi membrane when PAK4 is co-expressed with activated Cdc42. Current data strongly implicates PAK-4 in oncogenesis. PAK4 is frequently overexpressed in human tumor cell lines of various tissue origins.
Application Dilution
WB	1:500-2000
IHC-P	FCM1:100
IHC-F	IHC-P1:100-500
IF	ICC1:100
Flow-Cyt
ICC/IF