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<strong>Figure 1 KO Validation in HEK293T Cells</strong><br> Loading: 10 μg of HEK293T WT cell lysates or MANF KO cell lysates. Antibodies: MANF 4347 (1 μg/mL) and beta-actin 3779 (1 μg/mL), 1 h incubation at RT in 5% NFDM/TBST.Secondary: Goat Anti-Rabbit IgG HRP conjugate at 1:10000 dilution.
<strong>Figure 2 Western Blot Validation in Human, Mouse (NIH/3T3) and Rat (YB2/0) Cell Lines</strong><br>Loading: 15 μg of lysates per lane.Antibodies: MANF 4347, (2 μg /mL), 1h incubation at RT in 5% NFDM/TBST.Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.
<strong>Figure 3 Western Blot Validation with Recombinant Protein</strong><br>Loading: 30 ng of human MANF recombinant protein per lane.Antibodies: MANF 4347 (Lane 1: 0.125 μg /mL, Lane 2: 0.25μg /mL and Lane 3: 0.5 μg /mL), 1h incubation at RT in 5% NFDM/TBST.Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.Observed at around 55kD.
<strong>Figure 4 Western Blot Validation in Human Tissues </strong><br>Loading: 15 μg of lysates per lane.Antibodies: MANF 4347, (2 μg/mL), 1h incubation at RT in 5% NFDM/TBST.Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.
<strong>Figure 5 Western Blot Validation in Mouse Tissues </strong><br>Loading: 15 μg of lysates per lane.Antibodies: MANF 4347, (2 μg/mL), 1h incubation at RT in 5% NFDM/TBST.Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.
<strong>Figure 6 Western Blot Validation in Rat Liver </strong><br>Loading: 15 μg of lysates per lane.Antibodies: MANF 4347, 1h incubation at RT in 5% NFDM/TBST.Secondary: Goat anti-rabbit IgG HRP conjugate at 1:10000 dilution.Lane 1: 1 μg/mLLane 2: 2 μg/mLLane 3: 4 μg/mL
<strong>Figure 7 Immunofluorescence Validation of MANF in Human Cervix</strong><br>Immunofluorescent analysis of 4% paraformaldehyde-fixed human cervix cells labeling MANF with 4347 at 20 μg/mL, followed by goat anti-rabbit IgG secondary antibody at 1/500 dilution (green) and DAPI antibody (blue).
<strong>Figure 8 Immunofluorescence Validation of MANF in Human Colon </strong><br>Immunofluorescent analysis of 4% paraformaldehyde-fixed human colon cells labeling MANF with 4347 at 5 μg/mL, followed by goat anti-rabbit IgG secondary antibody at 1/500 dilution (green) and DAPI antibody (blue).
<strong>Figure 9 Immunofluorescence Validation of MANF in Human Liver </strong><br>Immunofluorescent analysis of 4% paraformaldehyde-fixed human liver cells labeling MANF with 4347 at 20 μg/mL, followed by goat anti-rabbit IgG secondary antibody at 1/500 dilution (green) and DAPI antibody (blue).
<strong>Figure 10 Immunofluorescence Validation of MANF in Mouse Pancreas </strong><br>Immunofluorescent analysis of 4% paraformaldehyde-fixed mouse pancreas cells labeling MANF with 4347 at 20 μg/mL, followed by goat anti-rabbit IgG secondary antibody at 1/500 dilution (green) and DAPI antibody (blue).
<strong>Figure 11 Immunofluorescence Validation of MANF in Rat Liver </strong><br>Immunofluorescent analysis of 4% paraformaldehyde-fixed rat liver cells labeling MANF with 4347 at 20 μg/mL, followed by goat anti-rabbit IgG secondary antibody at 1/500 dilution (green) and DAPI antibody (blue).
<strong>Figure 12 Immunohistochemistry Validation of MANF in Human Brain Tissue </strong><br> Immunohistochemical analysis of paraffin-embedded human brain tissue using anti-MANF antibody (4347) at 2.5 μg /ml. Tissue was fixed with formaldehyde and blocked with 10% serum for 1 h at RT; antigen retrieval was by heat mediation with a citrate buffer (pH6). Samples were incubated with primary antibody overnight at 4°C. A goat anti-rabbit IgG H&L (HRP) at 1/250 was used as secondary. Counter stained with Hematoxylin.
<strong>Figure 13 Localization and Immunocytochemistry Validation of MANF in adult zebrafish brain. (Chen et al., 2012) </strong><br>Co-immunostaining with anti-MANF antibodies (4347) shows that majority of MANF-positive cells were HuC/D positive cells (fig. C-E) whereas a few MANF-positive cells were either glial (zrf-1) positive cells (fig. F-H) or TH1 positive cells (fig I-K). It indicates that MANF was mostly expressed in mature neuronal cells. MANF staining is shown in green.
<strong>Figure 14 KD Validation of MANF in Zebrafish Embryo (Chen et al., 2012)</strong><br> Western blot analysis with anti-MANF antibodies (4347) was performed for MANF in 3dpf zebrafish embryo. After injected with a combination of two antisense splice-blocking morpholino oligonucleotides (MOsp), MANF expression was reduced by 85% as compared to the wild type.
<strong>Figure 15 Induced Expression Validation of MANF in Mouse C17.2 Neural Stem Cells (Almutawaa et al., 2014) </strong><br>Concentration- and time-dependent effects of Valproic acid (VPA) on MANF at 48 h examined by Western blot analysis with anti-MANF antibodies. MANF was markedly increased 48h after VPA treatment.Lanes 1–4: control, 0.5 mM VPA, 1 mM VPA, 3 mM VPA.
Figure 1 KO Validation in HEK293T Cells
Loading: 10 μg of HEK293T WT cell lysates or MANF KO cell lysates. Antibodies: MANF 4347 (1 μg/mL) and beta-actin 3779 (1 μg/mL), 1 h incubation at RT in 5% NFDM/TBST.Secondary: Goat Anti-Rabbit IgG HRP conjugate at 1:10000 dilution.
Cell Treatment
KO
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MANF Antibody

CATALOG NUMBER: 4347

Clonality:
Polyclonal
Tested Applications:
ELISA, IF, IHC-P, WB
Host Species:
Rabbit
Species Reactivity:
Human, Mouse, Rat
Conjugate:
Unconjugated
Specifications
Host Species:
Rabbit
Species Reactivity:
Human, Mouse, Rat
Homology:
Predicted species reactivity based on immunogen sequence: Bovine: (92%)
Immunogen:
MANF antibody was raised against a 12 amino acid synthetic peptide from near the carboxy terminus of human MANF.
The immunogen is located within the last 50 amino acids of MANF.
Conjugate:
Unconjugated
Tested Applications:
ELISA, IF, IHC-P, WB
Application Note:
WB: 0.125-2 μg/mL; IHC: 2.5 μg/mL; IF: 20 μg/mL
Antibody validated: Western Blot in human, mouse and rat samples; Immunohistochemistry in human samples; Immunofluorescence in human samples. All other applications and species not yet tested.
Specificity:
This antibody does not cross-react with CDNF.
Positive Control 1:
Cat. No. 1403 - Mouse Brain Tissue Lysate
Positive Control 2:
Cat. No. 10-301 - Human Brain Tissue Slide
Positive Control 3:
Cat. No. 1224 – Daudi Cell Lysate
Positive Control 4:
Cat. No. 1282 – 3T3/NIH Lysate
Predicted Molecular Weight:
Predicted: 20kD
Observed: 18 kD
Validation:

KO Validation (Figure 1): Anti-MANF antibodies (4347) specificity was further verified by MANF specific knockout. MANF signal was not detected in MANF knockout HEK293T cells as compared to that in control wild type cells.

Recombinant Protein Test (Figure 3): Anti-MANF antibodies (4347) detected human MANF recombinant protein at different concentrations.

Localization Validation (Figure 8): MANF detected by anti-MANF antibodies is colocalized with HuC/D (a general neural marker).  MANF is localized in mature neuronal cells.

KD Validation (Figure 9): Anti-MANF antibody (4347) specificity was further verified by MANF specific knockdown. MANF signal, when injected with a combination of two antisense splice-blocking morpholino oligonucleotides (MOsp), was disrupted in comparison with either wild type or when injected with control MO .

Induced Expression Validation (Figure 10): MANF expression detected by anit-MANF antibodies was up-regulated by VPA treatment.

Isoforms:
Human MANF has one isoform (182aa, 21kD). Mouse MANF has one isoform (179aa, 20kD) and Rat MANF also has one isoform (179aa, 20kD). 4347 can detect human, mouse and rat.
Purification:
MANF Antibody is Protein A purified.
Clonality:
Polyclonal
Isotype:
IgG
physical-state:
Liquid
Buffer:
MANF Antibody is supplied in PBS containing 0.02% sodium azide.
Concentration:
1 mg/mL
Storage Conditions:
MANF antibody can be stored at 4°C for three months and -20°C, stable for up to one year. As with all antibodies care should be taken to avoid repeated freeze thaw cycles. Antibodies should not be exposed to prolonged high temperatures.
Ncbi Official Symbol:
MANF
Additional Names:
MANF Antibody: ARP, ARMET, ARP, Mesencephalic astrocyte-derived neurotrophic factor, Arginine-rich protein
Protein Accession Number:
P55145
Protein Gi Number:
23503040
Ncbi Gene Id Number:
7873
User Note:
Optimal dilutions for each application to be determined by the researcher.
Background:
MANF Antibody: MANF, also known as ARMET, was initially identified as a protein containing an arginine-rich region that was highly mutated in a variety of tumors. More recently it was identified as a mesencephalic astrocyte-derived neurotrophic factor with selectivity for dopaminergic neurons, similar to glial cell line-derived neurotrophic factor (GDNF) and CDNF. In rat brain slices, MANF enhanced nigral gamma-aminobutyric acid release. Like GDNF and CDNF, MANF has selective neuroprotective activity for dopaminergic neurons suggesting that it may be indicated for the treatment of Parkinson's disease. Expression of MANF has also been shown to be induced during ER stress, suggesting that it may play a role in protein quality control during ER stress.
Background Reference 1:
Shridhar et al. Oncogene1996; 12:1931-9.
Background Reference 2:
Shridhar et al. Cancer Res.1996; 56:5576-8.
Background Reference 3:
Petrova et al. J. Mol. Neurosci.2003; 20:173-88.
Background Reference 4:
Lindholm et al. Nature 2007; 448:73-7.
Citation 1:
Chen et al. MANF regulates dopaminergic neuron development in larval zebrafish. Dev Biol. 2012;370(2):237-49. PMID: 22898306
Citation 2:
Almutawaa et al. Induction of neurotrophic and differentiation factors in neural stem cells by valproic acid. Basic Clin Pharmacol Toxicol. 2014;115(2):216-21. PMID: 24460582
Citation 3:
Niles et al. Valproic acid up-regulates melatonin MT1 and MT2 receptors and neurotrophic factors CDNF and MANF in the rat brain. Int J Neuropsychopharmacol. 2012;15(9):1343-50. PMID: 22243807
Citation 4:
Walaa S. Almutawaa. Induction Of Neurotrophic and Differentiation Genes in Neural Stem Cells by Valproic Acid. McMaster University, M.Sc. Thesis. 2012.PMID:
Citation 5:
Merja H. Voutilainen. CDNF and MANF in an experimental model of Parkinson's disease in rats. University of Helsinki, Ph.D. Thesis. 2010.PMID:

FOR RESEARCH USE ONLY

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Disclaimer:
Optimal dilutions/concentrations should be determined by the end user. The information provided is a guideline for product use. This product is for research use only.

CATALOG NUMBER:

4347

List Size:
0.02 mg, 0.1 mg

List Price:

Price range: $99.00 through $445.00

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