HMGB1 ELISA Kit

Catalog No. ABIN6574156

Product Details HMGB1 ELISA Kit

Target: HMGB1 (High Mobility Group Box 1 (HMGB1))

Reactivity: Mouse

Detection Method: Colorimetric

Method Type: Sandwich ELISA

Detection Range: 46.88 pg/mL - 3000 pg/mL

Minimum Detection Limit: 46.88 pg/mL

Application: ELISA

Purpose: The kit is a sandwich enzyme immunoassay for in vitro quantitative measurement of HMG1 in mouse serum, plasma.

We offer validation data (WB) for the kit components. So you can be sure to order a reliable ELISA kit product composed of high quality reagents.

Sample Type: Plasma, Serum

Analytical Method: Quantitative

Specificity: This assay has high sensitivity and excellent specificity for detection of High Mobility Group Protein 1 (HMGB1)

Cross-Reactivity (Details): No significant cross-reactivity or interference between High Mobility Group Protein 1 (HMG1) and analogues was observed.

Sensitivity: 18.29 pg/mL

Components:

• Pre-coated, ready to use 96-well strip plate, flat buttom
• Plate sealer for 96 wells
• Reference Standard
• Standard Diluent
• Detection Reagent A
• Detection Reagent B
• Assay Diluent A
• Assay Diluent B
• Reagent Diluent (if Detection Reagent is lyophilized)
• TMB Substrate
• Stop Solution
• Wash Buffer (30 x concentrate)
• Instruction manual

Application Details

Comment:

Information on standard material:
The standard might be recombinant protein or natural protein, that will depend on the specific kit. Moreover, the expression system is E.coli or yeast or mammal cell. There is 0.05% proclin 300 in the standard as preservative.

Information on reagents:
The stop solution used in the kit is sulfuric acid with concentration of 1 mol/L. And the wash solution is TBS. The standard diluent contains 0.02 % sodium azide, assay diluent A and assay diluent B contain 0.01% sodium azide. Some kits can contain is BSA in them.

Information on antibodies:
The provided antibodies and their host vary in different kits.

Sample Volume: 100 μL

Assay Time: 3 h

Plate: Pre-coated

Protocol:

1. Prepare all reagents, samples and standards,
2. Add 100μL standard or sample to each well. Incubate 1 hours at 37 °C,
3. Aspirate and add 100μL prepared Detection Reagent A. Incubate 1 hour at 37 °C,
4. Aspirate and wash 3 times,
5. Add 100μL prepared Detection Reagent B. Incubate 30 minutes at 37 °C,
6. Aspirate and wash 5 times,
7. Add 90μL Substrate Solution. Incubate 10-20 minutes at 37 °C,
8. Add 50μL Stop Solution. Read at 450nm immediately.

Reagent Preparation:

1. Bring all kit components and samples to room temperature (18-25 °C) before use. If the kit will not be used up in one time, please only take out strips and reagents for present experiment, and leave the remaining strips and reagents in required condition.
2. Standard - Reconstitute the Standard with 1.0 mL of Standard Diluent, keep for 10 minutes at room temperature, shake gently (not to foam). The concentration of the standard in the stock solution is 6,000pg/mL. Firstly dilute the stock solution to 3,000pg/mL and the diluted standard serves as the highest standard (3,000pg/mL). Then prepare 7 tubes containing 0.5 mL Standard Diluent and use the diluted standard to produce a double dilution series. Mix each tube thoroughly before the next transfer. Set up 7 points of diluted standard such as 3,000pg/mL, 1,500pg/mL, 750pg/mL, 375pg/mL, 187.5pg/mL, 93.75pg/mL, 46.88pg/mL, and the last microcentrifuge tube with Standard Diluent is the blank as 0pg/mL.
3. Detection Reagent A and Detection Reagent B - If lyophilized reconstitute the Detection Reagent A with 150μL of Reagent Diluent, keep for 10 minutes at room temperature, shake gently (not to foam). Briefly spin or centrifuge the stock Detection A and Detection B before use. Dilute them to the working concentration 100-fold with Assay Diluent A and B, respectively.
4. Wash Solution - Dilute 20 mL of Wash Solution concentrate (30x) with 580 mL of deionized or distilled water to prepare 600 mL of Wash Solution (1x).
5. TMB substrate - Aspirate the needed dosage of the solution with sterilized tips and do not dump the residual solution into the vial again.

Note:

1. Making serial dilution in the wells directly is not permitted.
2. Prepare standards within 15 minutes before assay. Please do not dissolve the reagents at 37 °C directly.
3. Please carefully reconstitute Standards or working Detection Reagent A and B according to the instruction, and avoid foaming and mix gently until the crystals are completely dissolved. To minimize imprecision caused by pipetting, use small volumes and ensure that pipettors are calibrated. It is recommended to suck more than 10μL for one pipetting.
4. The reconstituted Standards, Detection Reagent A and Detection Reagent B can be used only once.
5. If crystals have formed in the Wash Solution concentrate (30x), warm to room temperature and mix gently until the crystals are completely dissolved.
6. Contaminated water or container for reagent preparation will influence the detection result.

Sample Preparation:

• It is recommended to use fresh samples without long storage, otherwise protein degradation and denaturationmay occur in these samples, leading to false results. Samples should therefore be stored for a short periodat 2 - 8 °C or aliquoted at -20 °C (≤1 month) or -80 °C (≤ 3 months). Repeated freeze-thawcycles should be avoided. Prior to assay, the frozen samples should be slowly thawed and centrifuged toremove precipitates.
• If the sample type is not specified in the instructions, a preliminary test is necessary to determinecompatibility with the kit.
• If a lysis buffer is used to prepare tissue homogenates or cell culture supernatant, there is a possibilityof causing a deviation due to the introduced chemical substance.The recommended dilution factor is for reference only.
• Please estimate the concentration of the samples before performing the test. If the values are not in therange of the standard curve, the optimal sample dilution for the particular experiment has to be determined.Samples should then be diluted with PBS (pH =7.0-7.2).

Assay Precision: Intra-assay Precision (Precision within an assay): 3 samples with low, middle and high level of target were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, middle and high level of target were tested on 3 different plates, 8 replicates in each plate.
CV(%) = SD/meanX100
Intra-Assay: CV < 10%
Inter-Assay: CV < 12%

Restrictions: For Research Use only

Handling

Precaution of Use: The Stop Solution suggested for use with this kit is an acid solution. Wear eye, hand, face, and clothing protection when using this material.

Storage: 4 °C/-20 °C

Storage Comment:

1. For unopened kit: All reagents should be stored according to the labels on the vials. The Standard, Detection Reagent A, Detection Reagent B, and 96-well Strip Plate should be stored at -20 °C upon receipt, while the other reagents should be stored at 4 °C.
2. For opened kits: the remaining reagents must be stored according to the above storage conditions. In addition, please return the unused wells to the foil pouch containing the desiccant and seal the foil pouch with the zipper.

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Expiry Date: 6 months

References for HMGB1 ELISA Kit (ABIN6574156)

Lenkiewicz, Adamiak, Thapa, Bujko, Pedziwiatr, Abdel-Latif, Kucia, Ratajczak, Ratajczak: "The Nlrp3 Inflammasome Orchestrates Mobilization of Bone Marrow-Residing Stem Cells into Peripheral Blood." in: Stem cell reviews and reports, Vol. 15, Issue 3, pp. 391-403, (2019) (PubMed).

Lee, Wang, Li, Liu: "Anti-inflammatory effect of cinnamaldehyde and linalool from the leaf essential oil of Cinnamomum osmophloeum Kanehira in endotoxin-induced mice." in: Journal of food and drug analysis, Vol. 26, Issue 1, pp. 211-220, (2019) (PubMed).

Park, Kim, Kim, Chang: "Luteolin activates ERK1/2- and Ca2+-dependent HO-1 induction that reduces LPS-induced HMGB1, iNOS/NO, and COX-2 expression in RAW264.7 cells and mitigates acute lung injury of endotoxin mice." in: Inflammation research : official journal of the European Histamine Research Society ... [et al.], Vol. 67, Issue 5, pp. 445-453, (2018) (PubMed).

Kim, Park, Kim, Chang: "Sirt1 S-nitrosylation induces acetylation of HMGB1 in LPS-activated RAW264.7 cells and endotoxemic mice." in: Biochemical and biophysical research communications, Vol. 501, Issue 1, pp. 73-79, (2018) (PubMed).

Wang, Li, Deng, Liu, He: "Ursolic Acid Ameliorates Inflammation in Cerebral Ischemia and Reperfusion Injury Possibly via High Mobility Group Box 1/Toll-Like Receptor 4/NFκB Pathway." in: Frontiers in neurology, Vol. 9, pp. 253, (2018) (PubMed).

Peng, Liu, Ojcius, Lee, Chen, Huang, Martel, Young: "Mineral particles stimulate innate immunity through neutrophil extracellular traps containing HMGB1." in: Scientific reports, Vol. 7, Issue 1, pp. 16628, (2017) (PubMed).

Qian, Wei, Xu, He, Hua, Li, Hu, Lin, Gong, Meng, Zhou, Teng, Song: "Bone marrow-derived mesenchymal stem cells (BMSCs) repair acute necrotized pancreatitis by secreting microRNA-9 to target the NF-κB1/p50 gene in rats." in: Scientific reports, Vol. 7, Issue 1, pp. 581, (2017) (PubMed).

Chen, Fang, Li, Chen, Li, Gong, Fang: "Glycyrrhizin ameliorates experimental colitis through attenuating interleukin-17-producing T cell responses via regulating antigen-presenting cells." in: Immunologic research, Vol. 65, Issue 3, pp. 666-680, (2017) (PubMed).

Chaochao, Lou, Yang, Liu, Hu, Min, Chen, He, Chen: "Macrophage Inflammatory Protein-2 in High Mobility Group Box 1 Secretion of Macrophage Cells Exposed to Lipopolysaccharide." in: Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology, Vol. 42, Issue 3, pp. 913-928, (2017) (PubMed).

Yadav, Rani, Deep, Singh, Palle: "Oxidative Stress in Metabolic Disorders: Pathogenesis, Prevention, and Therapeutics." in: Oxidative medicine and cellular longevity, Vol. 2016, pp. 9137629, (2016) (PubMed).

Yu, Yu, Liu, Yu, Liu, Liu, Su, Jiang, Chen: "Ethyl pyruvate attenuated coxsackievirus B3-induced acute viral myocarditis by suppression of HMGB1/RAGE/NF-ΚB pathway." in: SpringerPlus, Vol. 5, pp. 215, (2016) (PubMed).

Kopecka, Porto, Lusa, Gazzano, Salzano, Pinzòn-Daza, Giordano, Desiderio, Ghigo, De Rosa, Caraglia, Riganti: "Zoledronic acid-encapsulating self-assembling nanoparticles and doxorubicin: a combinatorial approach to overcome simultaneously chemoresistance and immunoresistance in breast tumors." in: Oncotarget, Vol. 7, Issue 15, pp. 20753-72, (2016) (PubMed).

Liu, Ma, Sun, Li, Wang: "High Mobility Group Box1 Protein Is Involved in Endoplasmic Reticulum Stress Induced by Clostridium difficile Toxin A." in: BioMed research international, Vol. 2016, pp. 4130834, (2016) (PubMed).

Chen, Li, Khan, Shi, Wang, Zheng, Gong, Fang: "HMGB1 exacerbates experimental mouse colitis by enhancing innate lymphoid cells 3 inflammatory responses via promoted IL-23 production." in: Innate immunity, Vol. 22, Issue 8, pp. 696-705, (2016) (PubMed).

Wu, Sheng, Xie, Li, Chen, Li, Wang, Xu: "Reduced HMGB 1-Mediated Pathway and Oxidative Stress in Resveratrol-Treated Diabetic Mice: A Possible Mechanism of Cardioprotection of Resveratrol in Diabetes Mellitus." in: Oxidative medicine and cellular longevity, Vol. 2016, pp. 9836860, (2016) (PubMed).

Kim, Ha, Kim, Park, Kim, Park, Kim, Chung, Chang: "Ascorbic acid reduces HMGB1 secretion in lipopolysaccharide-activated RAW 264.7 cells and improves survival rate in septic mice by activation of Nrf2/HO-1 signals." in: Biochemical pharmacology, Vol. 95, Issue 4, pp. 279-89, (2015) (PubMed).

Sun, Chen, Dai, Zou, Gao, Wu, Ming, Lai, Xiao, Xiong, Xu, Gong, Zheng: "HMGB1 expression patterns during the progression of experimental autoimmune encephalomyelitis." in: Journal of neuroimmunology, Vol. 280, pp. 29-35, (2015) (PubMed).

Kim, Kim, Chang: "Glycyrrhizin reduces HMGB1 secretion in lipopolysaccharide-activated RAW 264.7 cells and endotoxemic mice by p38/Nrf2-dependent induction of HO-1." in: International immunopharmacology, Vol. 26, Issue 1, pp. 112-8, (2015) (PubMed).

Chen, Sun, Lai, Wu, Xiao, Ming, Gao, Zou, Xiong, Xu, Tan, Gong, Zheng: "Interleukin-33 is released in spinal cord and suppresses experimental autoimmune encephalomyelitis in mice." in: Neuroscience, Vol. 308, pp. 157-68, (2015) (PubMed).

Jiang, Wang, Yun, Hajrasouliha, Zhao, Sun, Kaplan, Shao: "HMGB1 release triggered by the interaction of live retinal cells and uveitogenic T cells is Fas/FasL activation-dependent." in: Journal of neuroinflammation, Vol. 12, pp. 179, (2015) (PubMed).

Target Details for HMGB1

Target: HMGB1 (High Mobility Group Box 1 (HMGB1))

Alternative Name: High Mobility Group Protein 1 (HMGB1) (HMGB1 Products)

Synonyms: HMG1 ELISA Kit, HMG3 ELISA Kit, SBP-1 ELISA Kit, DEF ELISA Kit, HMG-1 ELISA Kit, Hmg1 ELISA Kit, amphoterin ELISA Kit, p30 ELISA Kit, hmgb1 ELISA Kit, ik:tdsubc_1a5 ELISA Kit, wu:fb23c02 ELISA Kit, xx:tdsubc_1a5 ELISA Kit, zgc:56110 ELISA Kit, zgc:77104 ELISA Kit, hmg-1 ELISA Kit, hmg3 ELISA Kit, sbp-1 ELISA Kit, hmg1 ELISA Kit, HMGB1 ELISA Kit, Ac2-008 ELISA Kit, high mobility group box 1 ELISA Kit, high-mobility group box 1 ELISA Kit, high mobility group box 1a ELISA Kit, high mobility group box 1 L homeolog ELISA Kit, high mobility group protein B1 ELISA Kit, HMGB1 ELISA Kit, Hmgb1 ELISA Kit, hmgb1 ELISA Kit, hmgb1a ELISA Kit, hmgb1.L ELISA Kit, LOC100359149 ELISA Kit

UniProt: P63158

Pathways: p53 Signaling, Regulation of Muscle Cell Differentiation, Skeletal Muscle Fiber Development, Positive Regulation of Endopeptidase Activity, Regulation of Carbohydrate Metabolic Process, Toll-Like Receptors Cascades, Smooth Muscle Cell Migration, Inflammasome