Human tight junction protein 1 (ZO1) ELISA Kit

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Target Background

Product Details

Target Name

tight junction protein 1 (zona occludens 1)

Alternative Names

Tight junction protein 1 ELISA Kit; Tight junction protein ZO-1 ELISA Kit; Tight junction protein ZO1 ELISA Kit; TJP1 ELISA Kit; zo-1 ELISA Kit; Zo1 ELISA Kit; ZO1_HUMAN ELISA Kit; Zona occludens 1 ELISA Kit; Zona occludens 1 protein ELISA Kit; Zona occludens protein 1 ELISA Kit; Zonula occludens 1 protein ELISA Kit; Zonula occludens protein 1 ELISA Kit

Abbreviation

TJP1

Uniprot No.

Q07157

Species

Homo sapiens (Human)

Sample Types

serum, plasma, cell culture supernates, cell lysates

Detection Range

15.6 pg/mL-1000 pg/mL

Sensitivity

3.9 pg/mL

Assay Time

1-5h

Sample Volume

50-100ul

Detection Wavelength

450 nm

Research Area

Tags & Cell Markers

Assay Principle

quantitative

Measurement

Sandwich

Precision

Linearity

To assess the linearity of the assay, samples were spiked with high concentrations of human ZO1 in various matrices and diluted with the Sample Diluent to produce samples with values within the dynamic range of the assay.
	Sample	Serum(n=4)
1:1	Average %	84
	Range %	80-90
1:2	Average %	99
	Range %	91-104
1:4	Average %	100
	Range %	92-108
1:8	Average %	94
	Range %	86-98

Recovery

The recovery of human ZO1 spiked to levels throughout the range of the assay in various matrices was evaluated. Samples were diluted prior to assay as directed in the Sample Preparation section.

Sample Type	Average % Recovery	Range
Serum (n=5)	95	89-97
EDTA plasma (n=4)	97	90-99

Typical Data

These standard curves are provided for demonstration only. A standard curve should be generated for each set of samples assayed.

pg/ml	OD1	OD2	Average	Corrected
1000	2.484	2.467	2.476	2.329
500	1.707	1.678	1.693	1.546
250	1.076	1.055	1.066	0.919
125	0.679	0.669	0.674	0.527
62.5	0.441	0.436	0.439	0.292
31.2	0.297	0.287	0.292	0.145
15.6	0.222	0.209	0.216	0.069
0	0.149	0.145	0.147

Materials provided

A micro ELISA plate --- The 96-well plate has been pre-coated with an anti-human ZO-1 antibody. This dismountable microplate can be divided into 12 x 8 strip plates.
Two vials lyophilized standard ---Dilute a bottle of the standard at dilution series, read the OD values, and then draw a standard curve.
One vial Biotin-labeled ZO-1 antibody (100 x concentrate) (120 μl/bottle) ---Act as the detection antibody.
One vial HRP-avidin (100 x concentrate) (120 μl/bottle) ---Bind to the detection antibody and react with the TMB substrate to make the solution chromogenic.
One vial Biotin-antibody Diluent (15 ml/bottle) ---Dilute the Biotin-antibody.
One vial HRP-avidin Diluent (15 ml/bottle) ---Dilute the HRP-avidin solution.
One vial Sample Diluent (50 ml/bottle)---Dilute the sample to an appropriate concentration.
One vial Wash Buffer (25 x concentrate) (20 ml/bottle) ---Wash away unbound or free substances.
One vial TMB Substrate (10 ml/bottle) ---Act as the chromogenic agent. TMB interacts with HRP, eliciting the solution turns blue.
One vial Stop Solution (10 ml/bottle) ---Stop the color reaction. The solution color immediately turns from blue to yellow.
Four Adhesive Strips (For 96 wells) --- Cover the microplate when incubation.
An instruction manual

Materials not provided

A microplate reader capable of measuring absorbance at 450 nm, with the correction wavelength set at 540 nm or 570 nm.
An incubator can provide stable incubation conditions up to 37°C±5°C.
Centrifuge
Vortex
Squirt bottle, manifold dispenser, or automated microplate washer
Absorbent paper for blotting the microtiter plate
50-300ul multi-channel micropipette
Pipette tips
Single-channel micropipette with different ranges
100ml and 500ml graduated cylinders
Deionized or distilled water
Timer
Test tubes for dilution

ELISA Data Analysis

ELISA Standard Curve & Curve Expert software

Troubleshooting
and FAQs

ELISA kit FAQs

Storage

Store at 2-8°C. Please refer to protocol.

Lead Time

3-5 working days after you place the order, and it takes another 3-5 days for delivery via DHL or FedEx

Description

The Human tight junction protein 1 (ZO-1) ELISA kit is a sandwich immunoassay specifically designed and validated for the quantitative detection of ZO-1 in the serum, plasma, cell culture supernates, or cell lysates. It is not intended for diagnostic use. This assay kit was designed and optimized for Tags & Cell Markers research use in humans. The kit has undergone rigorous quality control in multiple parameters, including sensitivity, specificity, precision, linearity, recovery, and inter-batch difference. Refer to the product instructions for more details. This assay employs the quantitative sandwich enzyme immunoassay technique, in which ZO-1 in the samples or standards are sandwiched between pre-coated ZO-1 antibody and Biotin-conjugated antibody specific for ZO-1. HRP-avidin is then added to the wells. Following a wash to remove any unbound reagent, the TMB substrate solution is added to the wells and color develops in proportion to the amount of ZO-1 bound in the initial step. The color development is stopped upon adding the stop solution, and the intensity of the color is measured at 450 nm via a microplate reader. The levels of ZO-1 in the samples are determined by comparing the O.D. (optical density) of the samples to the standard curve.

ZO-1, also called TJP1, is a tight junction (TJ) phosphoprotein acting as a scaffold protein that cross-links and anchors tight junction strand proteins to the actin cytoskeleton. ZO-1 organizes and regulates the structure of tight junctions through direct interactions with transmembrane factors including claudin (CLDN), occludin (OCLN), and junction adhesion molecule A (JAM-A). Besides, ZO-1 shuttles between the nucleus and plasma membrane modulate paracellular permeability and participate in the regulation of gene expression. ZO-1 can also bind to ZONAB through its Src homology 3 (SH3) domain. ZONAB is involved in the control of epithelial cell proliferation. ZO1 was down-regulated in many cancers and decreased ZO1 expression may be closely related to patient prognosis.

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Target Background

Function
(From Uniprot)

TJP1, TJP2, and TJP3 are closely related scaffolding proteins that link tight junction (TJ) transmembrane proteins such as claudins, junctional adhesion molecules, and occludin to the actin cytoskeleton. The tight junction acts to limit movement of substances through the paracellular space and as a boundary between the compositionally distinct apical and basolateral plasma membrane domains of epithelial and endothelial cells. Necessary for lumenogenesis, and particularly efficient epithelial polarization and barrier formation. Plays a role in the regulation of cell migration by targeting CDC42BPB to the leading edge of migrating cells. Plays an important role in podosome formation and associated function, thus regulating cell adhesion and matrix remodeling. With TJP2 and TJP3, participates in the junctional retention and stability of the transcription factor DBPA, but is not involved in its shuttling to the nucleus.

Gene References into Functions

Dysfunction of the miR-455-TJP1 axis is involved in bladder cancer cell growth and metastasis. PMID: 30061227
The tight junction protein ZO-1 exists in stretched and folded conformations within epithelial cells, depending on actomyosin-generated force. PMID: 29199076
miR103 was upregulated in CRC. Overexpression of miR103 promoted CRC cell proliferation and migration in vitro, whereas downregulation of miR103 inhibited cell proliferation and migration. ZO1 was identified as a direct target of miR103, revealing its expression to be inversely correlated with miR103 expression in CRC samples. PMID: 29115525
SHANK3 expression correlated with ZO-1 and PKCepsilon in colonic tissue of patients with Crohn's disease. The expression level of SHANK3 affects ZO-1 expression and the barrier function in intestinal epithelial cells. PMID: 28906292
These results indicate the varying effects of 7-oxygenated cholesterol molecules on the expression and localization of ZO-1 depending on cell types, and suggest the contribution of 7-oxygeneted cholesterol molecules to the structural alteration of tight junctions. PMID: 29428726
CTR activates AKAP2-anchored cAMP-dependent protein kinase A, which then phosphorylates tight junction proteins ZO-1 and claudin 3. PMID: 28428082
the Ras signaling pathway is involved in HIV-1 Tat-induced changes in ZO-1 and NEP. PMID: 28553432
decreased interaction between ZO-1 and occludin might contribute to the epiphora occurred in the transplanted submandibular glands PMID: 28332063
integration of claudin-2, occludin and ZO-1 is necessary for maintaining the function of the proximal tubular epithelium. PMID: 29252987
Endothelial cellsTLR4 strongly regulates retinal vessel permeability by reducing expression of occludin and zonula occludens 1. PMID: 29136627
The role of estrogens in the regulation of ZO-1 and estrogen receptors 1 and 2 was evaluated in human primary gut tissues by immunohistochemistry, immunofluorescence and qPCR. PMID: 28867253
Aberrant expression of the tight junction molecules claudin-1 and zonula occludens-1 mediates cell growth and invasion in oral squamous cell carcinoma cells. PMID: 27436828
ZO-1-occludin interactions regulate multiple phases of epithelial polarization by providing cell-intrinsic signals that are required for single lumen formation. PMID: 27802160
It is postulated that ZO-1, when not phosphorylated by PKC, keeps Octn2 in an active state, while elimination of this binding in DeltaPDZ mutant or after ZO-1 phosphorylation leads to diminution of Octn2 activity. PMID: 28257821
Results uncovered ZO-1 as part of a signaling node activated by VEGF, but not Ang-1, that specifically modulates endothelial cells proliferation during angiogenesis. PMID: 26846344
Data suggest that long noncoding RNA PlncRNA1 and microRNA miR-34c bound together to regulate the expressions of MAZ, ZO-1 and occludin. PMID: 28153728
ZO-1 highly expresses in cell-cell junctions and is related to odontoblast differentiation, which may contribute to dental pulp repair or even the formation of an odontoblast layer. PMID: 27109589
this study shows that the expression and the immunoreactivity of ZO-1 is decreased in the nasal epithelium of patients with allergic rhinitis PMID: 27216347
The frequency of alleles and genotypes of rs2291166 gene polymorphism TJP1 was determined in the Mexico Mestizos population. The ancestral allele was the most prevalent. The conformational effect of this amino acid change was performed in silico. PMID: 26259745
We identified potential nuclear and membrane biomarkers (increased expression of ZO-1, caveolin-1 and P2X7 receptor) of risks for placenta and pregnancy PMID: 26657896
OCLN and ZO1 levels appear to be early prognostic markers in patients suffering from sepsis. PMID: 26863122
The data suggest that ZO-1, along with CD38 and Zap-70, plays a role in cell cycle regulation in chronic B cell leukemia, and may be used as a prognostic marker in the disease monitoring. PMID: 26306999
These data provide the first evidence that beta-catenin and ZO-1 are direct targets of E7 of the oncogenic beta-human papillomavirus types 5 and 8. PMID: 26645068
Report TNF-alpha/Il6 mediated dysregulation of zonula occludens-1 properties in human brain microvascular endothelium. PMID: 25953589
upon specific knockdown of the accessory TJP, ZO-1, undifferentiated NSCs showed decreased levels of key stem cell markers PMID: 25892136
HTT may inhibit breast tumor dissemination through maintenance of ZO1 at tight junctions. PMID: 26293574
These results suggest that the localization of ZO-1 in cell-cell contacts is differently regulated by activation and inhibition of JNK and/or p38 MAPK depending on the incubation period. PMID: 25435485
CFTR colocalizes with ZO-1 at the tight junctions of trachea and epididymis, and is expressed before ZO-1 in Wolffian ducts. PMID: 25107366
Phosphorylation state of the tyrosine of claudin-1 and claudin-2 regulates interaction with ZO1. PMID: 26023235
miR-18a and RUNX1 could reversely regulate the permeability of blood-tumor barrier as well as the expressions and distributions of ZO-1, occludin and claudin-5. PMID: 25452107
Zonula occludens-1, occludin and E-cadherin expression and organization in salivary glands PMID: 25248927
ZO-1 was internalized and shown to accumulate in the cytoplasm of human podocytes in an IL-13 dose-dependent manner. PMID: 25683991
The decreased UCP2 expression and increased ZO-1 expression suggest that the oxidative stress-induced mitochondrial dysfunction and tight junction formation may play pivotal roles in the progress of NVG. PMID: 23835672
ZO-1 is a central regulator of VE-cadherin-dependent endothelial junctions that orchestrates the spatial actomyosin organization. PMID: 25753039
Tjp1 expression was decreased in glomerular diseases in human and animal models, our results indicate that the suppression of Tjp1 could directly aggravate glomerular disorders, highlights Tjp1 as a potential therapeutic target. PMID: 25184792
In conclusion, our present study indicated that miR-34c regulated the permeability of BTB via MAZ-mediated expression changes of ZO-1, occludin, and claudin-5. PMID: 25201524
ZO-1 showed a tendency to be detected more intensely in myocardial infarction and ischemic heart disease myocardial tissue then in asphyxiation or drowning. PMID: 24368520
ZO-1 gene shows a hypermethylation status in children with NHL. PMID: 24927439
gene expression is regulated by p38MAPK PMID: 23856837
High expression of ZO-1 is associated with good prognosis in non-small cell lung cancer. PMID: 24294375
The LIM domain protein FHL1C interacts with tight junction protein ZO-1 contributing to the epithelial-mesenchymal transition of a breast adenocarcinoma cell line. PMID: 24657059
luciferase assays and chromatin immunoprecipitation assays showed that KLF4 up-regulated the promoter activities and interacted with "CACCC" DNA sequence presented in the promoters of ZO-1, occludin, and claudin-5. PMID: 24318462
our results identify a novel regulatory pathway involving the interplay between ZO-1, alpha5-integrin and PKCepsilon in the late stages of mammalian cell division. PMID: 23967087
Data suggest components of diet supplements (here, glutamine/arginine) can improve permeability and tight junction protein expression (TJP1/occludin) in enterocytes exposed to deleterious effects of antineoplastic agents (here, methotrexate). PMID: 23428392
ZO-1 expression is correlated with malignant phenotypes of GIST. PMID: 23820955
Vascular endothelial tight junctions and barrier function are disrupted by 15(S)-hydroxyeicosatetraenoic acid partly via protein kinase C epsilon-mediated zona occludens-1 phosphorylation at threonine 770/772. PMID: 24338688
The presence of neural cells (PC12 cells or trigeminal neurons) markedly promoted the stratification of HCE cells as well as increased the amounts of N-cadherin mRNA and protein in these cells. PMID: 24327615
The methylation positivity rates of the ID4 and ZO-1 genes in the bone marrow and paraffin-embedded lymphoma tissues of non-Hodgkin lymphoma patients were significantly higher compared to the rates in the Hodgkin lymphoma patients. PMID: 23670122
Proteomic identification of ZO-1 binding partners and associated proteins that form tight junction complexes. PMID: 23553632
Single nucleotide polymorphisms in TJP1 is associated with response to antipsychotic agents in schizophrenia. PMID: 23241943

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Subcellular Location

Cell membrane; Peripheral membrane protein; Cytoplasmic side. Cell junction, tight junction. Cell junction. Cell junction, gap junction. Cell projection, podosome.

Protein Families

MAGUK family

Tissue Specificity

The alpha-containing isoform is found in most epithelial cell junctions. The short isoform is found both in endothelial cells and the highly specialized epithelial junctions of renal glomeruli and Sertoli cells of the seminiferous tubules.

Database Links

HGNC: 11827

OMIM: 601009

KEGG: hsa:7082

STRING: 9606.ENSP00000281537

UniGene: Hs.743990

Code	CSB-E13916h
Size	96T,5×96T,10×96T
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Intra-assay Precision (Precision within an assay): CV%<8%

Three samples of known concentration were tested twenty times on one plate to assess.

Inter-assay Precision (Precision between assays): CV%<10%

Three samples of known concentration were tested in twenty assays to assess.