40-3400

antibody from Invitrogen Antibodies

Targeting: GRN CLN11, PCDGF, PGRN

Provider product page for 40-3400

Western blot

Immunoprecipitation

Other assay

Antibody data

Antibody Data
Antigen structure
References [13]
Comments [0]
Validations
Western blot [1]
Other assay [17]

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Product number: 40-3400 - Provider product page
Provider: Invitrogen Antibodies
Product name: PCDGF Polyclonal Antibody
Antibody type: Polyclonal
Antigen: Synthetic peptide
Reactivity: Human
Host: Rabbit
Isotype: IgG
Vial size: 100 µg
Concentration: 0.25 mg/mL
Storage: -20°C

GRN protein structure - 40-3400 shown in red.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: NULL

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 1 HIV infection induces PGRN production in microglia. Primary human microglial cells were inoculated with VSVg env HIV or HIV ADA and PGRN in culture supernatants were determined by ELISA as described in the Materials and Methods. Control cultures were treated with VSVg env protein or mock infected. (A) Results with VSVg env HIV are shown. Culture supernatants were collected at 2 D and 4 D with complete change of medium (mean +- SD from triplicate cultures) (B) Microglia were incubated with AZT (10 ug/ml) or vehicle for 1 h, then exposed to VSVg env HIV as in A. PGRN was measured at 7 D and 14 D with complete change of medium. Data shown are accumulation between 7D-14D (mean +- SD, n = 3). (C) HIV ADA (HIV env bearing virus) was used to determine PGRN production as described in the Materials and Methods. Data shown are PGRN accumulation in culture supernatants between 7D-14D (mean +- SD, n = 3) (D) Microglial cultures infected with VSVg env HIV in B showing cell viability, gag p24 expression, and complete suppression of p24 by AZT treatment. **P

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 2 PGRN is an endogenous anti-HIV factor. Microglial PGRN was knocked down using RNAi 2-4 days prior to VSVg env HIV exposure as described in the Materials and Methods. Control cultures were treated with control, irrelevant siRNA (Ctr). The amounts of HIV (p24) and PGRN expression were determined by western blot analyses. (A) A representative western blot showing suppression of PGRN and increase of p24 following PGRN siRNA treatment. (B) Pooled densitometry data from four independent experiments showing significant inhibition of PGRN and increase of HIV ( gag p24 express) in microglial cells treated with PGRN siRNA (vs. control siRNA). **P

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig. 1 STAT3 interacts with GRN in breast cancer cells (A) Mass spectrometry coverage of GRN amino acid sequence P28799. Peptides identified by PEAKS or Proteome Discoverer are indicated in red underline. (B) STAT3-GRN interaction in MDA-MB-468 and SUM159PT cells was analyzed by immunoprecipitation using antibodies against STAT3 (or non-specific immunoglobulin G, IgG) followed by immunoblots with the indicated antibodies. Input indicates 5% of pre-immunoprecipitated samples.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig. 2 GRN is necessary for constitutive STAT3 transcriptional activity in triple-negative breast cancer cells MDA-MB-468 and SUM159PT cells were transfected with siRNA targeting GRN (pool siGRN and individual constructs siGRN-B and siGRN-C) or a non-targeting control (siCon). They were then analyzed by (A) immunoblot for levels of phosphorylated and total STAT3 in whole cell lysates, (B) luciferase reporter assay for STAT3-dependent transcriptional activity (N = 3), (C) nuclear and cytoplasmic fractions (tubulin serves as a loading control for the cytoplasmic fraction and PARP serves as a loading control for the nuclear fraction), (D) qRT-PCR for expression of endogenous STAT3 target genes (normalized to GAPDH; N = 3), and (E) chromatin immunoprecipitation with an antibody for STAT3 followed by PCR at the indicated STAT3 target genes (representative of N = 3).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig. 3 GRN is necessary for maximal cytokine-induced STAT3 transcriptional activity in breast cancercells (A) STAT3-GRN interaction in SK-BR-3 cells was analyzed by immunoprecipitation using antibodies against STAT3 (or non-specific IgG) followed by immunoblot with the indicated antibodies. Input indicates 5% of pre-immunoprecipitated samples. SK-BR-3 cells were transfected with siRNA targeting GRN (pool siGRN and individual constructs siGRN-B and siGRN-C) or a non-targeting control (siCon), then stimulated with LIF were analyzed by (B) immunoblot and (C) luciferase reporter assay for STAT3-dependent transcriptional activity. SK-BR-3 cells transfected with siRNA targeting GRN (pool) were analyzed by (D) luciferase reporter assay for STAT3-dependent transcriptional activity (B-luc, M67-luc) (N = 3), (E) qRT-PCR for expression of endogenous STAT3 target genes (normalized to HPRT; N = 6), and (F) chromatin immunoprecipitation with an antibody to STAT3 followed by PCR at the indicated STAT3 target genes (data normalized to unstimulated cells; representative of N = 2).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Fig. 4 GRN enhances the kinetics of cytokine-stimulated STAT3 activation and nuclear translocation (A) SK-BR-3 cells were transfected with siRNA targeting GRN or a non-targeting control, then stimulated with LIF for the indicated times. Cells were then analyzed by immunoblot for levels of phosphorylated and total STAT3 in whole cell extracts. Tubulin serves as a loading control. (B) Immunoblot band intensities plotted as ratios of phosphorylated STAT3 to tubulin. (C) Area under the curve of phospho-STAT3:tubulin (N = 5). (D) SK-BR-3 cells were transfected with siRNA targeting GRN or a non-targeting control, then stimulated with LIF for the indicated times. Cells were then analyzed by immunoblot for levels of phosphorylated and total STAT3 in nuclear fractions. PARP serves as a nuclear fraction loading control. (E) Immunoblot band intensities plotted as ratios of phospho-STAT3 to PARP (N = 4).

Supportive validation

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Experimental details: FIGURE 1 Recombinant Progranulin Protein Adsorbs to Polypropylene Tubes. (A,B) 100 nM rhPGRN was serially transferred through tubes 1-5, which were stripped for adsorbed protein after a 10-min incubation in each tube. Tube 6 contains the protein remaining in solution at the end of five serial transfers. The control is 100 nM PGRN that underwent no transfers. Low binding pipet tips were used to minimize pipetting loss. One-Way ANOVA with Holm-Sidak's Multiple Comparisons test was performed (**** p < 0.0001, 1-6 vs. Control, n = 3). (C) A dilution series of 250-25 nM PGRN was set up in polypropylene tubes. After a 10-min incubation on ice, the PGRN in solution as well as adsorbed to every tube was prepared for analysis. Low binding pipet tips were used to minimize pipetting loss. (D) 100 nM PGRN was prepared in tubes 1-6. Polypropylene tips were used to aspirate and dispense the full volume of solution 0-5 times, respectively, into the same tube. The control is 100 nM PGRN subjected to no pipetting. PGRN levels were measured by Western blot and membranes were immunoblotted with anti-PGRN antibody. All immunoblots are representative of three independent experiments.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: FIGURE 2 Time and Temperature Impact Adsorption of Recombinant Progranulin to Polypropylene Tubes. (A) 100 nM PGRN was incubated for 10', 1, 4, 8, 16, 24, or 48 h in polypropylene tubes on ice. One-Way ANOVA with Dunnett's Multiple Comparisons were performed (* p < 0.05, 1/4/8/16/24/48 h vs. 10', n = 3). (B) 100 nM PGRN was incubated in polypropylene tubes for 10' on ice, at room temperature (RT) or at 37degC ( n = 3). At the end of the incubations, PGRN in solution as well as adsorbed to the tube was prepared for analysis. PGRN levels were measured by Western blot and membranes were immunoblotted with anti-PGRN antibody. All immunoblots are representative of three independent experiments.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: FIGURE 3 PGRN Adsorption is Prevented in BSA-Coated Tubes. (A) A dilution series of 250-25 nM PGRN was set up in BSA-coated polypropylene tubes for 10-min incubations on ice ( n = 3). (B) 100 nM PGRN was incubated for 10', 1, 4, 8, 16, 24, or 48 h in BSA-coated tubes on ice ( n = 3). (C) 100 nM PGRN was incubated in BSA-coated tubes for 10' on ice, at room temperature (RT) or at 37degC ( n = 3). At the end of the incubations, PGRN in solution as well as adsorbed to the tube was prepared for analysis. PGRN levels were measured by Western blot and membranes were immunoblotted with anti-PGRN antibody. All immunoblots are representative of three independent experiments.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: FIGURE 4 PGRN Adsorption is Reduced in LoBind Tubes. (A) A dilution series of 250-25 nM PGRN was set up in LoBind tubes for 10-min incubations on ice ( n = 3). (B) 100 nM PGRN was incubated for 10', 1, 4, 8, 16, 24 or 48 h in LoBind tubes on ice ( n = 3). (C) 100 nM PGRN was incubated in LoBind tubes for 10' on ice, at room temperature (RT) or at 37degC ( n = 3). At the end of the incubations, PGRN in solution as well as adsorbed to the tube was prepared for analysis. PGRN levels were measured by Western blot and membranes were immunoblotted with anti-PGRN antibody. All immunoblots are representative of three independent experiments.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Figure 1. Expression of recombinant human GRNs and identification of antibodies that detect GRNs. A , Schematic of human GRN expression constructs. Human GRN sequences (Table 2), with and without adjacent C-terminal linker regions, were synthesized to include the N-terminal PGRN signal peptide (SP), followed by twin-Strep (SAWSHPQFEK) tags and a single FLAG (DYKDDDDK) tag. Throughout the manuscript, individual GRNs are referred to by their numerical sequential designation (i.e., GRN-1, GRN-2, etc.), which correspond to their original alphabetical designation (i.e., GRN-G, GRN-F, etc.). B , C , HEK293T cells were transfected with the human GRN constructs and 48 h later either ( B ) whole-cell lysates or ( C ) conditioned media were analyzed by immunoblot for protein expression using the StrepMAB-immo antibody. D , Cell lysates from GRN(+linker) overexpressing cells were mock (-) or PNGase F (+) treated to detect glycosylated proteins. E , F , GRN-expressing HEK293T cell lysates were analyzed by immunoblot to identify PGRN antibodies that either ( E ) detect single linker regions of PGRN or ( F ) detect GRNs. The specific PGRN linker or GRN(s) detected are listed under each antibody. Detailed information about the PGRN antibodies screened can be found in Table 3. Asterisks (*) denote nonspecific protein bands.