- 701056 - Invitrogen Antibodies MAPT antibody

Submitted references Ageing and amyloidosis underlie the molecular and pathological alterations of tau in a mouse model of familial Alzheimer's disease.

Metaxas A, Thygesen C, Kempf SJ, Anzalone M, Vaitheeswaran R, Petersen S, Landau AM, Audrain H, Teeling JL, Darvesh S, Brooks DJ, Larsen MR, Finsen B
Scientific reports 2019 Oct 31;9(1):15758

Early Evidence of Low Bone Density and Decreased Serotonergic Synthesis in the Dorsal Raphe of a Tauopathy Model of Alzheimer's Disease.

Dengler-Crish CM, Smith MA, Wilson GN
Journal of Alzheimer's disease : JAD 2017;55(4):1605-1619

Effects of antibodies to phosphorylated and non-phosphorylated tau on in vitro tau phosphorylation at Serine-199: Preliminary report.

Loeffler DA, Smith LM, Klaver AC, Martić S
Experimental gerontology 2015 Jul;67:15-8

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of Phospho-Tau pThr231 in total rat brain lysate using a Phospho-Tau pThr231 recombinant rabbit monoclonal antibody (Product # 701056) at a dilution of 1 µg/mL. To confirm specificity, competition was performed by preincubation with the phosphopeptide to inhibit antibody binding (lane 2). Samples were detected using chemiluminescence (ECL). Results show a band at ~56kDa.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of Phospho-Tau pThr231 in total rat brain lysate using a Phospho-Tau pThr231 recombinant rabbit monoclonal antibody (Product # 701056) at a dilution of 1 µg/mL. To confirm specificity, competition was performed by preincubation with the phosphopeptide to inhibit antibody binding (lane 2). Samples were detected using chemiluminescence (ECL). Results show a band at ~56kDa.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of Tau (pT231) was performed by loading 30 µg of rat brain lysate (lane 1) using Novex®NuPAGE®4-12% Bis-Tris gel (Product # NP0321BOX), XCell SureLock Electrophoresis System (Product # EI0002), Novex® Sharp Pre-Stained Protein Standard (Product # LC5800), and iBlot® Dry Blotting System (Product # IB21001). Proteins were transferred to a nitrocellulose membrane and blocked with 5% skim milk for 1 hour at room temperature. Tau (pT231) was detected at ~55 kDa using Tau (pT231) Recombinant Rabbit Monoclonal Antibody (Product # 701056) at a 1:1000 dilution in 2.5% skim milk at 4°C overnight on a rocking platform. To confirm specificity, competition was performed with the phosphopeptide (10 µg/mL) (lane 2). Detection was performed using an HRP-conjugated Goat anti-Rabbit secondary antibody (Product # G-21234) at a 1:5000 dilution and chemiluminescent detection was performed using Pierce™ ECL Western blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunofluorescent analysis of Tau (pT231) was performed on 70% confluent log phase HeLa cells. The cells were fixed with 4% paraformaldehyde for 15 minutes, permeabilized with 0. 25% Triton X-100 for 10 minutes, and blocked with 5% BSA for 1 hour at room temperature. The cells were labeled with Tau (pT231) Recombinant Rabbit Monoclonal Antibody (Product # 701056) at a dilution of 1:500 in 1% BSA and incubated for 3 hours at room temperature and then labeled with Alexa Fluor® 488 Goat anti-Rabbit IgG secondary antibody (Product # A-11008) at a dilution of 1:400 for 30 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® 594 phalloidin (Product # A12381). Panel d is a merged image showing nuclear localization and panel e shows competition with the phospho-Tau (pT231) peptide.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of Phospho-Tau pThr231 showing staining in the cytoplasm of paraffin-embedded human brain tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10 mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with Phospho-Tau pThr231 (1H6L8) Monoclonal antibody (Product # 701056) diluted in 3% BSA-PBS at a dilution of 1:200 overnight at 4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using a HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of Phospho-Tau pThr231 showing staining in the cytoplasm of paraffin-embedded human glioma (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10 mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a Phospho-Tau pThr231 (1H6L8) Monoclonal antibody (Product # 701056) diluted in 3% BSA-PBS at a dilution of 1:200 overnight at 4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using a HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of Phospho-Tau pThr231 showing staining in the cytoplasm of paraffin-embedded rat brain tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10 mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a Phospho-Tau pThr231 (1H6L8) Monoclonal antibody (Product # 701056) diluted in 3% BSA-PBS at a dilution of 1:200 overnight at 4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using a HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig.4 Phosphorylated tau (ptau) accumulation in trytophan hydroxylase (TPH)-positive cells of the DRN is evident in 4-month-old htau mice. Photomicrographs of immunofluorescent-stained DRN sections are shown in the first three columns for C57, tau -/- , and htau mice. All mice exhibit distinctly-labeled TPH cells; however, ptau-231 label is non-existent in C57 and tau -/- DRN. ptau-231 immunofluorescence is obvious in DRN sections of 4-month-old htau mice with substantial co-label of this marker in TPH-positive cells. Fourth column: Photomicrographs of merged channels of TPH and ptau for MRN from each respective animal. Note that ptau-231 fluorescence in htau MRN is weak compared to DRN. Scale bar = 100 mum.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Fig.5 Co-localization of ptau-231 in tryptophan hydroxylase (TPH)-positive cells of 4-month-old htau DRN. A) Confocal images of single planes through rostral DRN at the level of the trochlear nuclei for C57, tau -/- , and htau mice. Phosphorylated tau (ptau) fluorescence is co-localized to TPH-positive cells in the htau section only. Scale bar = 10 mum. B) 3-D reconstruction of a TPH-positive cell in the htau DRN. Orthogonal views of the stack depict fluorescence label across tissue depth. Crosshairs (dashed lines) indicate the plane of section for the X, Y, and Z dimensions. The TPH-positive cell shows prominent ptau label in the cytoplasm of the soma with sparser puncta in TPH-positive processes.Scale bar = 10 mum.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Figure 6 Validation of pS404, pT231 and 3R tau. ( A) Immunoblotting of sarkosyl-insoluble tau with rabbit primary antibody against phospho-Ser404 (1:200; OAAF07796, Aviva Systems Biology). The entire membrane is shown. Hyperphosphorylation at the S404 residue was exclusively observed in 24-month-old APP swe /PS1 DeltaE9 mice (TG 24 M, lane 8). Lanes are labelled as follows: Marker: 1, 14; AD: 3; non-AD: 5; WT 24 months: 6; TG 24 months: 8; WT 3 months: 10; TG 3 months: 12; Empty: 2, 4, 7, 9, 11, 13. (B) ELISA of pT231 tau. Soluble pT231 tau was present in the neocortex of both WT and TG mice. Phosphorylation at T231 was only observed in the sarkosyl-insoluble fraction from 24-month-old APP swe /PS1 DeltaE9 mice. Results are expressed as arbitrary units (U), normalized to total protein concentration. (C) Immunohistochemistry of pT231 tau in coronal, 50 um-thick brain sections from 3- and 18-month-old WT and TG mice. Black arrows point to CA1 pyramidal neurons, which were more strongly immunolabelled in 18 vs. 3-month-old animals, irrespective of genotype. Scale bar: 200 um. The inserts show higher magnifications of layer V neurons in the temporal cortex, with black arrowheads pointing to tangle-like structures in aged APP swe /PS1 DeltaE9 mice. Note reduced dendritic staining in 18 vs. 3-month-old animals. Scale bar: 10 um. White arrows point to puncta of pT231 immunoreactivity within a plaque-like structure, observed exclusively in the neocortex of TG mice. Scale b

701056

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