PA1-1745

antibody from Invitrogen Antibodies

Targeting: ACAN AGC1, CSPG1, CSPGCP, MSK16

Western blot (Supportive data in Antibodypedia)

Immunocytochemistry (Supportive data in Antibodypedia)

Antibody Data

Product number

PA1-1745

Provider

Invitrogen Antibodies

Product name

Anti-Aggrecan G3 Polyclonal Antibody

Antibody type

Polyclonal

Antigen

Synthetic peptide

Description

PA1-1745 detects human, rat, mouse, bovine, and canine aggrecan G3. PA1-1745 has been successfully used in Western blot procedures. By Western blot, this antibody detects an ~350 kDa protein representing the aggrecan G3 domain on the full length core. The PA1-1745 immunogen is a synthetic peptide corresponding to residues C D(2277) G H P M Q F E N W R P N Q P D N(2293) of human aggrecan.. This antibody is also referred to as anti-G3.

Reactivity

Human, Mouse, Rat, Bovine, Canine

Host

Rabbit

Isotype

IgG

Vial size

100 µg

Concentration

1 mg/ml

Storage

-20° C, Avoid Freeze/Thaw Cycles

References

Submitted references

Biochemical and atomic force microscopic characterization of salmon nasal cartilage proteoglycan.

Kakizaki I, Mineta T, Sasaki M, Tatara Y, Makino E, Kato Y
Carbohydrate polymers 2014 Mar 15;103:538-49

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MMP proteolysis of the human extracellular matrix protein aggrecan is mainly a process of normal turnover.

Struglics A, Hansson M
The Biochemical journal 2012 Sep 1;446(2):213-23

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Aggrecanase cleavage in juvenile idiopathic arthritis patients is minimally detected in the aggrecan interglobular domain but robust at the aggrecan C-terminus.

Struglics A, Lohmander LS, Last K, Akikusa J, Allen R, Fosang AJ
Arthritis and rheumatism 2012 Dec;64(12):4151-61; author reply 4162-3

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Identification of proteoglycan from salmon nasal cartilage.

Kakizaki I, Tatara Y, Majima M, Kato Y, Endo M
Archives of biochemistry and biophysics 2011 Feb 1;506(1):58-65

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Calpain is involved in C-terminal truncation of human aggrecan.

Struglics A, Hansson M
The Biochemical journal 2010 Sep 15;430(3):531-8

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A comparison of different purification methods of aggrecan fragments from human articular cartilage and synovial fluid.

Struglics A, Larsson S
Matrix biology : journal of the International Society for Matrix Biology 2010 Jan;29(1):74-83

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Figure 1 illustrates Western blot detection of mouse chondrocyte cells using PA1-1745. The specificity of PA1-1745 is demonstrated in lane 1. Lane 1: mouse chondrocyte Lane 2: a lane of human cartilage proteoglycan blotted with anti-aggrecan G1 domain (general)

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot analysis was performed on whole cell extracts (30 µg lysate) of NIH/3T3 (Lane 1). The blots were probed with Anti-Aggrecan G3 alpha Rabbit Polyclonal Antibody (Product # PA1-1745, 1-3 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). A 239 kDa band corresponding to Aggrecan G3 was observed in the cell line tested. A lower band of 137 kDa was observed showing the proteolytic fragment of Aggrecan G3 and a higher band of ~300 kDa is a core protein which has around 100 GAG (glycosaminoglycans) chains attached to it. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 10% Bis-Tris gel (Product # NP0302BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and HiMark™ Pre-stained Protein Standard (Product # LC5699). Resolved proteins were then transferred onto a nitrocellulose membrane with overnight wet transfer System. The membrane was probed with the relevant primary and secondary Antibody following blocking with 5% skimmed milk. Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Immunocytochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunofluorescence analysis of Aggrecan G3 was performed using 70% confluent log phase A549 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 Aggrecan G3 Rabbit Polyclonal Antibody (Product # PA1-1745) 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 localization in the extracellular matrix. Panel e shows the no primary antibody control. The images were captured at 60X magnification.