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Khb0041

Manufactured by Thermo Fisher Scientific
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The KHB0041 is a laboratory equipment product from Thermo Fisher Scientific. It is designed for specific laboratory applications. The core function of this product is to perform tasks related to sample preparation, analysis, or other laboratory procedures. However, a detailed description of the product's features and capabilities cannot be provided in an unbiased and factual manner without the risk of extrapolation or interpretation.

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Lab products found in correlation

Khb7031, by Thermo Fisher Scientific (5 mentions) Khb8051, by Thermo Fisher Scientific (4 mentions) Kho0631, by Thermo Fisher Scientific (3 mentions) Dy401, by R&D Systems (2 mentions) Khb3481, by Thermo Fisher Scientific (2 mentions) Khb3544, by Thermo Fisher Scientific (2 mentions) Infinite m200 pro, by Tecan (1 mentions) Sodium orthovanadate, by Merck Group (1 mentions) Triton x 100, by Merck Group (1 mentions) Bradford assay, by Bio-Rad (1 mentions) Phenyl methyl sulfonyl fluoride (pmsf), by Merck Group (1 mentions) Bio plex 200 system, by Bio-Rad (1 mentions) Protein lobind tube, by Eppendorf (1 mentions) Tau total human elisa kit, by Thermo Fisher Scientific (1 mentions) Multiskan fc, by Thermo Fisher Scientific (1 mentions) Khb7041, by Thermo Fisher Scientific (1 mentions) Versamax, by Molecular Devices (1 mentions) P tau s396, by Thermo Fisher Scientific (1 mentions) Mouse specific total tau elisa kit, by Thermo Fisher Scientific (1 mentions) Ps199 tau elisa kit, by Thermo Fisher Scientific (1 mentions)

20 protocols using khb0041

1

Extracellular Tau Quantification in Cortisol-Treated Cells

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Cell cultures were treated with 2 μM Cortisol plus 200 nM DFP, and their supernatant was collected with protease inhibitors (Thermo Scientific, 78430) and phosphatase inhibitors (Thermo Scientific, 78428) and analyzed fresh for extracellular human total tau (Invitrogen, KHB0041) and [pT231] human phosphorylated tau (Invitrogen, KHB8051) according to the manufacturer’s instructions. The absorbance was read on a Tecan Infinite M200Pro spectral plate reader for optical density with subsequent analyses to calculate the concentration of extracellular human total tau or [pT231] human phosphorylated tau in each sample. See supplemental material for more detailed methods.
Yates P.L., Patil A., Sun X., Niceforo A., Gill R., Callahan P., Beck W., Piermarini E., Terry A.V., Sullivan K.A., Baas P.W, & Qiang L. (2021). A cellular approach to understanding and treating Gulf War Illness. Cellular and molecular life sciences : CMLS, 78(21-22), 6941-6961.
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2

Quantification of Tau Protein and IL-1beta

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Total tau and pT231 tau contents were measured by commercial tau ELISA kits according to the manufacturer’s instructions (total tau - KHB0041; pT231 tau - KHB8051, Invitrogen), as previously described (Swarup et al., 2019 (link)). Mouse IL-1beta was quantified using cell culture supernatant via ELISA (R&D Systems, DY401) according to the manufacturer’s protocol, with absorbance measured at 450nm.
Rexach J.E., Polioudakis D., Yin A., Swarup V., Chang T.S., Nguyen T., Sarkar A., Chen L., Huang J., Lin L.C., Seeley W., Trojanowski J.Q., Malhotra D, & Geschwind D.H. (2020). Tau Pathology Drives Dementia Risk-Associated Gene Networks toward Chronic Inflammatory States and Immunosuppression. Cell reports, 33(7), 108398.
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3

Quantifying Total and Phospho-Tau in Cell Lysates

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Total tau and phosphorylated (p) tau (Threonine 181, T181) were analyzed by ELISA assay (KHB0041 and KHO0631, respectively; Invitrogen; Italy) on cell lysates. Briefly, cell lysates were obtained using a specific lysis buffer prepared adding to RIPA Buffer 1X protease inhibitor cocktail (Roche; Italy), 1% Triton X-100 (Sigma), 1% sodium orthovanadate (Sigma; UK), and 1% PMSF (Sigma). Cell lysates were quantified by the Bradford assay (Biorad; Italy), and then, 20 μg of the total lysate was diluted in 50 or 100 μl of the specific ELISA diluent and seeded into pre-coated wells. The presence of both total tau and ptau (T181) in the samples was revealed by colorimetric reaction and read at 450 nm, and concentration determined through interpolation to standard curve and reported as picograms per milliliter.
Bortolotti D., Gentili V., Rotola A., Caselli E, & Rizzo R. (2019). HHV-6A infection induces amyloid-beta expression and activation of microglial cells. Alzheimer's Research & Therapy, 11, 104.
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4

Quantifying Alzheimer's Protein Secretion

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To analyze protein secretion under the serum-deprived condition, the culture medium was changed to Opti-MEM and samples were incubated at 37 °C under 5% CO2 for 12 h. The culture medium was collected and centrifuged at 3000g for 10 min at 4 °C, and the supernatant was collected and stored at −80 °C. The proteins levels of Aβ1-40 (27713, IBL), Aβ1-42 (27711, IBL), phospho-tau (pT181) (KHO0631, Invitrogen), and total-tau (KHB0041, Invitrogen) were measured by enzyme-linked immunosorbent assay (ELISA) according to manufacturer’s instructions. BCA analysis was performed with the same samples, and the data were normalized by the total protein content. Levels of Aβs secreted from E3par and E4iso iCOs were further measured using xMAP technology (Bioplex 200 systems). The utilized protocol is also described in our previous paper66 (link).
Park J.C., Jang S.Y., Lee D., Lee J., Kang U., Chang H., Kim H.J., Han S.H., Seo J., Choi M., Lee D.Y., Byun M.S., Yi D., Cho K.H, & Mook-Jung I. (2021). A logical network-based drug-screening platform for Alzheimer’s disease representing pathological features of human brain organoids. Nature Communications, 12, 280.
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5

Quantification of Tau Protein and IL-1beta

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Total tau and pT231 tau contents were measured by commercial tau ELISA kits according to the manufacturer’s instructions (total tau - KHB0041; pT231 tau - KHB8051, Invitrogen), as previously described (Swarup et al., 2019 (link)). Mouse IL-1beta was quantified using cell culture supernatant via ELISA (R&D Systems, DY401) according to the manufacturer’s protocol, with absorbance measured at 450nm.
Rexach J.E., Polioudakis D., Yin A., Swarup V., Chang T.S., Nguyen T., Sarkar A., Chen L., Huang J., Lin L.C., Seeley W., Trojanowski J.Q., Malhotra D, & Geschwind D.H. (2020). Tau Pathology Drives Dementia Risk-Associated Gene Networks toward Chronic Inflammatory States and Immunosuppression. Cell reports, 33(7), 108398.
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6

Quantification of Alzheimer's Biomarkers

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We dissolved Aβ42 (1 mg) and Aβ40 (1 mg) in 480 μL of 1,1,1,3,3,3-hexafluoro-2-propanol and kept them overnight at room temperature. The solution was dispensed into Protein Lobind tube (Eppendorf AG, Germany) and vacuum-dried for 2 h. We dissolved human tau (100 μg) and synthetic p-tau181 (1 mg) in the deionized water. The solutions were aliquoted and stored at −20 °C for further experiments. We prepared biomarkers of femtomolar to picomolar concentrations by serially diluting the stock solution of each biomarker using a PBS solution (10 mM, pH 7.4). The stock concentration of each biomarker was 100 μg mL−1. We quantified the concentrations of biomarkers using an enzyme-linked immunosorbent assay (Invitrogen, Catalog Number KHB0041, Total Tau Human ELISA Kit) (Supplementary Fig. 35). We applied each biomarker to the bioreceptor-immobilized CNT device and incubated them for 15 min at the room temperature. For detecting of AD biomarker in human plasma, the blood plasma was diluted into 1/10. The resistance of the sensor chip was measured using a digital multimeter (Fluke 83 V, Fluke Co., USA).
Kim K., Kim M.J., Kim D.W., Kim S.Y., Park S, & Park C.B. (2020). Clinically accurate diagnosis of Alzheimer’s disease via multiplexed sensing of core biomarkers in human plasma. Nature Communications, 11, 119.
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7

Amyloid-β Solubility and Tau Quantification

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Amyloid-β (Aβ) solubility was determined in soluble fractions from, (1) 0.2% diethylamine (DEA), (2) radioimmuniprecipitation assay buffer (RIPA), and (3) 70% formic acid (FA), using sequential isolation conditions, respectively, as previously described (Murphy et al, 2007; Niedowicz et al, 2013; 2014). Aβ was quantified with a sandwich ELISA, as previously described (Murphy et al, 2007; Niedowicz et al, 2013; 2014). Aβ40 and Aβ42 were measured in the DEA and RIPA fractions using commercially available kits (Invitrogen, KHB3481 and KHB3544, respectively), according to the manufacturer’s instructions. To detect total Aβ in the FA fraction, capture antibody Ab42.5 (against Aβ1-16; 0.5 μg / well) was used, followed by detection antibody: 0.25 μg/mL of biotinylated 4G8 (against Aβ17-24; BioLegend, San Diego, CA), as we had not previously tested the commercial kits using FA. Aβ levels were normalized to total protein concentrations loaded per well, as determined by bicinchoninic acid assay (Pierce). Phosphorylated tau pT181 (KHO0631) and total tau (KHB0041) was measured in the RIPA fraction (Invitrogen). Personnel performing the assays were blinded to mouse treatment condition.
Duncan M.J., Guerriero L.E., Kohler K., Beechem L.E., Gillis B.D., Salisbury F., Wessel C., Wang J., Sunderam S., Bachstetter A.D., O’Hara B.F, & Murphy M.P. (2021). Chronic fragmentation of the daily sleep-wake rhythm increases amyloid-beta levels and neuroinflammation in the 3xTg-AD mouse model of Alzheimer’s disease. Neuroscience, 481, 111-122.
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8

Quantifying Alzheimer's Biomarkers in Plasma

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Plasma contents of total tau, APOE, amyloid beta-peptide 1-40, and amyloid beta-peptide 1-42 were measured using commercial ELISA kits according to the manufacturer's instructions (Total Tau, KHB0041, Invitrogen; APOE, ELH-ApoE4-1, RayBiotech; Amyloid beta 1-40, RE59781, IBL, and Amyloid beta 1-42, KHB3544, Thermo Fisher Scientific). Briefly, standard assays for detecting radioimmunoprecipitation assay-soluble samples were applied to the ELISA plates. After washing, a biotin-conjugated detection antibody was applied. Then, the positive reaction was enhanced with streptavidin–horseradish peroxidase and colored by 3,3′,5,5′-tetramethylbenzidine. The absorbance at 450 nm was applied, and the concentrations of four different proteins were calculated from the standard curves. All measurements were carried out in one round of experiments, and the results were read on a microplate photometer (MultiskanTM FC, Thermo Fisher).
Yang J., Wu S., Yang J., Zhang Q, & Dong X. (2023). Amyloid beta-correlated plasma metabolite dysregulation in Alzheimer's disease: an untargeted metabolism exploration using high-resolution mass spectrometry toward future clinical diagnosis. Frontiers in Aging Neuroscience, 15, 1189659.
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9

Quantifying Tau Protein Levels in Synaptic Fractions

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ELISA kits were used to detect levels of total human tau (catalog #KHB0041, Invitrogen) and human tau phosphorylated at S199 (catalog #KHB7041, Invitrogen), T231 (catalog #KHB8051, Invitrogen), and S396 (catalog #KHB7031, Invitrogen). For synaptosomal total tau, samples were diluted 1:20 (1 µg protein per well) and the calculated concentration was multiplied by 20 to account for the dilution. For insoluble total tau, samples were diluted 1:1000 (0.02 µg protein per well) and the calculated concentration was multiplied by 1000 to account for the dilution. For all phospho-tau ELISAs, 20 µg of protein were loaded per well. ELISAs were read at 450 nm on a VersaMax microplate reader (Molecular Devices).
Walker C.K., Greathouse K.M., Tuscher J.J., Dammer E.B., Weber A.J., Liu E., Curtis K.A., Boros B.D., Freeman C.D., Seo J.V., Ramdas R., Hurst C., Duong D.M., Gearing M., Murchison C.F., Day J.J., Seyfried N.T, & Herskowitz J.H. (2023). Cross-Platform Synaptic Network Analysis of Human Entorhinal Cortex Identifies TWF2 as a Modulator of Dendritic Spine Length. The Journal of Neuroscience, 43(20), 3764-3785.
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10

Measuring Hippocampal tau Levels

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The mice were anesthetized with 2% isoflurane and subjected to a stereotaxic surgery to insert a cannula into dentate gyrus (DG). The method of microdialysis in vivo was described previously [16 (link)]. An ELISA kit was applied to measure total hTau present in microdialysis samples according to manufacturer’s instructions (KHB0041, Thermo Fisher).
Liu Y., Zhang S., Li X., Liu E., Wang X., Zhou Q., Ye J, & Wang J.Z. (2020). Peripheral inflammation promotes brain tau transmission via disrupting blood–brain barrier. Bioscience Reports, 40(2), BSR20193629.
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