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Human Zonulin ELISA Kit

Citations(37)Uniprot : P00738

  • Cat.No.:E-EL-H5560

  • Reactivity: Human
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  • 96T
  • 48T
  • 24T
  • 96T*5
  • 96T*10
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Product Details

Properties

Assay type Sandwich-ELISA
Format 96T/48T
Assay time 3.5h
Detection range 0.78-50 ng/mL
Sensitivity 0.47 ng/mL
Sample type &Sample volume serum, plasma and other biological fluids; 100μL
Specificity This kit recognizes Human Zonulin in samples. No significant cross-reactivity or interference between Human Zonulin and analogues was observed.
Reproducibility Both intra-CV and inter-CV are < 10%.
Application This ELISA kit applies to the in vitro quantitative determination of Human Zonulin concentrations in serum, plasma and other biological fluids.

Test Principle

This ELISA kit uses the Sandwich-ELISA principle. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Human Zonulin. Standards or samples are added to the micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human Zonulin and Avidin-Horseradish Peroxidase (HRP) conjugate are added successively to each micro plate well and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human Zonulin, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by the addition of stop solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Human Zonulin. You can calculate the concentration of Human Zonulin in the samples by comparing the OD of the samples to the standard curve.

Kit components & Storage

An unopened kit can be stored at 2-8℃ for 1 month. If the kit is not supposed to be used within 1 month, store the components separately according to the following conditions once the kit is received.
ItemSpecificationsStorage
Micro ELISA Plate(Dismountable) 96T: 8 wells ×12 strips
48T: 8 wells ×6 strips 		-20℃, 6 months
Reference Standard 96T: 2 vials
48T: 1 vial
Concentrated Biotinylated Detection Ab (100×) 96T: 1 vial, 120 μL
48T: 1 vial, 60 μL
Concentrated HRP Conjugate (100×) 96T: 1 vial, 120 μL
48T: 1 vial, 60 μL 		-20℃(Protect from light), 6 months
Reference Standard & Sample Diluent 1 vial, 20 mL 2-8°C, 6 months
Biotinylated Detection Ab Diluent 1 vial, 14 mL
HRP Conjugate Diluent 1 vial, 14 mL
Concentrated Wash Buffer (25×) 1 vial, 30 mL
Substrate Reagent 1 vial, 10 mL 2-8℃(Protect from light)
Stop Solution 1 vial, 10 mL 2-8°C
Plate Sealer 5 pieces
Manual 1 copy
Certificate of Analysis 1 copy

Technical Data

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human Zonulin were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human Zonulin were tested on 3 different plates, 20 replicates in each plate.

  Intra-assay Precision Inter-assay Precision
Sample 1 2 3 1 2 3
n 20 20 20 20 20 20
Mean (ng/mL) 2.46 5.57 22.64 2.31 6.06 23.45
Standard deviation 0.14 0.27 1.23 0.12 0.28 1.02
CV (%) 5.69 4.85 5.43 5.19 4.62 4.35

Recovery

The recovery of Human Zonulin spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.

Sample Type Range (%) Average Recovery (%)
Serum(n=8) 91-103 95
EDTA plasma (n=8) 87-99 91
Cell culture media (n=8) 91-99 95

Linearity

Samples were spiked with high concentrations of Human Zonulin and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.

Target Information

Database LinksSwissProt:   P00738
Research AreaCancer, Cardiovascular

Assay Procedures

elisa assay procedure 1

1. Add 100μL standard or sample to the wells. Incubate for 90 min at 37°C
elisa assay procedure 2

2. Discard the liquid, immediately add 100μL Biotinylated Detection Ab working solution to each well. Incubate for 60 min at 37°C
elisa assay procedure 3

3. Aspirate and wash the plate for 3 times
elisa assay procedure 4

4. Add 100μL HRP conjugate working solution. Incubate for 30 min at 37°C. Aspirate and wash the plate for 5 times
elisa assay procedure 5

5. Add 90μL Substrate Reagent. Incubate for 15 min at 37°C
elisa assay procedure 6

6. Add 50μL Stop Solution
elisa assay procedure 7

7. Read the plate at 450nm immediately. Calculation of the results

Citations

  1. HEPATOLOGY (2020) IF: 17.425
    Increased Liver Localization of Lipopolysaccharides in Human and Experimental NAFLD

    DOI: 10.1002/hep.31056

    PMID: 31808577

    		 Sample: Serum
  2. ANTIOXIDANTS & REDOX SIGNALING (2020) IF: 7.04
    Is There an Association Between Atherosclerotic Burden, Oxidative Stress, and Gut-Derived Lipopolysaccharides?

    DOI: 10.1089/ars.2020.8109
    Sample: serum
  3. FOOD CHEMISTRY (2021) IF: 7.514
    In vitro digestibility and Caco-2 cell bioavailability of sea lettuce (Ulva fenestrata) proteins extracted using pH-shift processing

    DOI: 10.1016/j.foodchem.2021.129683

    PMID: 33845254

    		 Sample: Cell
  4. Frontiers in Immunology (2021) IF: 7.561
    Detection of Alterations in the Gut Microbiota and Intestinal Permeability in Patients With Hashimoto Thyroiditis

    DOI: 10.3389/fimmu.2021.579140
    Sample: serum
  5. INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES (2022) IF: 6.208
    Macrophage Activation Markers Predict Liver-Related Complications in Primary Biliary Cholangitis

    DOI: 10.3390/ijms23179814

    PMID: 36077228

    		 Sample: serum
  6. Nutrients (2022) IF: 6.706
    Effects of Flavonoid-Rich Orange Juice Intervention on Major Depressive Disorder in Young Adults: A Randomized Controlled Trial

    DOI: 10.3390/nu15010145
    Sample: serum
  7. Oxidative Medicine and Cellular Longevity (2020) IF: 5.076
    Oxidative Stress and Gut-Derived Lipopolysaccharides in Neurodegenerative Disease: Role of NOX2

    DOI: 10.1155/2020/8630275

    PMID: 32089785

    		 Sample: Serum
  8. Nutrients (2021) IF: 5.719
    Microbiome as Mediator of Diet on Colorectal Cancer Risk: The Role of Vitamin D, Markers of Inflammation and Adipokines

    DOI: 10.3390/nu13020363

    PMID: 33504116

    		 Sample: Serum
  9. Nutrients (2022) IF: 5.719
    Extra Virgin Olive Oil Reduces Gut Permeability and Metabolic Endotoxemia in Diabetic Patients

    DOI: 10.3390/nu14102153

    PMID: 35631294

    		 Sample: serum
  10. ANTIOXIDANTS & REDOX SIGNALING (2019) IF: 5.828
    Does Nox2 Overactivate in Children with Nonalcoholic Fatty Liver Disease?

    DOI: 10.1089/ars.2018.7596
    Sample: Serum
  11. PHARMACOLOGICAL RESEARCH (2020) IF: 5.893
    Probiotics and fructo-oligosaccharide intervention modulate the microbiota-gut brain axis to improve autism spectrum reducing also the hyper-serotonergic state and the dopamine metabolism disorder

    DOI: 10.1016/j.phrs.2020.104784
    Sample: plasma
  12. INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES (2021) IF: 5.924
    Examining the Gastrointestinal and Immunomodulatory Effects of the Novel Probiotic Bacillus subtilis DE111

    DOI: 10.3390/ijms22052453

    PMID: 33671071

    		 Sample: Plasma
  13. Scientific Reports (2018) IF: 4.122
    Localization of lipopolysaccharide from Escherichia Coli into human atherosclerotic plaque

    DOI: 10.1038/s41598-018-22076-4
    Sample: Serum
  14. AIDS (2022) IF: 4.177
    Plasma d-amino acids are associated with markers of immune activation and organ dysfunction in people with HIV

    DOI: 10.1097/QAD.0000000000003207

    PMID: 35212669

    		 Sample: plasma
  15. Clinical and Translational Gastroenterology (2021) IF: 4.488
    Low-Grade Endotoxemia and Thrombosis in COVID-19

    DOI: 10.14309/ctg.0000000000000348

    PMID: 34092777

    		 Sample: serum
  16. Nutrients (2020) IF: 4.546
    Intestinal Permeability in Children with Celiac Disease after the Administration of Oligofructose-Enriched Inulin into a Gluten-Free Diet—Results of a Randomized, Placebo-Controlled, Pilot Trial

    DOI: 10.3390/nu12061736
    Sample: Plasma
  17. Biomolecules (2019) IF: 4.694
    Plasma Zonulin Levels as a Non-Invasive Biomarker of Intestinal Permeability in Women with Gestational Diabetes Mellitus

    DOI: 10.3390/biom9010024

    PMID: 30641999

    		 Sample: Plasma
  18. Biomedicines (2023) IF: 4.757
    The Effects of Probiotics on Small Intestinal Microbiota Composition, Inflammatory Cytokines and Intestinal Permeability in Patients with Non-Alcoholic Fatty Liver Disease

    DOI: 10.3390/biomedicines11020640
    Sample: serum
  19. HIV MEDICINE (2021) IF: 3.18
    Lipopolysaccharide induces platelet activation in HIV patients: the role of different viral load patterns

    DOI: 10.1111/hiv.13059

    PMID: 33426758

    		 Sample: Serum
  20. ENDOCRINE (2020) IF: 3.235
    Could there be a role of serum zonulin increase in the development of hypercalcemia in primary hyperparathyroidism

    DOI: 10.1007/s12020-020-02504-0

    PMID: 32989570

    		 Sample: Serum
  21. EUROPEAN ARCHIVES OF PSYCHIATRY AND CLINICAL NEUROSCIENCE (2020) IF: 3.288
    Serum zonulin and claudin-5 levels in patients with schizophrenia

    DOI: 10.1007/s00406-020-01152-9
    Sample: serum
  22. International Journal of Environmental Research and Public Health (2021) IF: 3.39
    Detection of Dysbiosis and Increased Intestinal Permeability in Brazilian Patients with Relapsing–Remitting Multiple Sclerosis

    DOI: 10.3390/ijerph18094621
    Sample: Serum
  23. JOURNAL OF MOLECULAR NEUROSCIENCE (2022) IF: 3.444
    Investigation of the Relation between Epithelial Barrier Function and Autism Symptom Severity in Children with Autism Spectrum Disorder

    DOI: 10.1007/s12031-021-01954-z

    PMID: 34988901

    		 Sample: serum
  24. NUTRITION METABOLISM AND CARDIOVASCULAR DISEASES (2017) IF: 3.679
    Low-grade endotoxemia, gut permeability and platelet activation in patients with impaired fasting glucose

    DOI: 10.1016/j.numecd.2017.06.007
    Sample: serum
  25. Scientific Reports (2020) IF: 3.998
    Assessment of haptoglobin alleles in autism spectrum disorders

    DOI: 10.1038/s41598-020-64679-w

    PMID: 32385356

    		 Sample: Serum
  26. INTERNATIONAL JOURNAL OF PSYCHIATRY IN CLINICAL PRACTICE (2020) IF: 2.144
    Serum zonulin and claudin-5 levels in children with attention-deficit/hyperactivity disorder

    DOI: 10.1080/13651501.2020.1801754

    PMID: 32757874

    		 Sample: Serum
  27. ARCHIVES OF GYNECOLOGY AND OBSTETRICS (2019) IF: 2.199
    Serum zonulin level is not elevated in patients with polycystic ovary syndrome without metabolic syndrome

    DOI: 10.1007/s00404-019-05345-z
    Sample: serum
  28. Journal of Cosmetic Dermatology (2022) IF: 2.696
    Is there a link between alopecia areata and gut?

    DOI: 10.1111/jocd.15095

    PMID: 35579378

    		 Sample: serum
  29. CHRONOBIOLOGY INTERNATIONAL (2022) IF: 2.877
    Changes in intestinal parameters and their association with dietary patterns in rotational shift workers

    DOI: 10.1080/07420528.2022.2044349

    PMID: 35232303

    		 Sample: Serum
  30. NORDIC JOURNAL OF PSYCHIATRY (2020) IF: 1.78
    Serum zonulin and claudin-5 levels in children with obsessive–compulsive disorder

    DOI: 10.1080/08039488.2020.1715474
    Sample: serum
  31. Journal of Clinical Research in Pediatric Endocrinology (2020) IF: 1.803
    Children with Hashimoto’s Thyroiditis Have Increased Intestinal Permeability: Results of a Pilot Study

    DOI: 10.4274/jcrpe.galenos.2020.2019.0186

    PMID: 31990165

    		 Sample: serum
  32. BMC Pediatrics (2020) IF: 1.909
    Oxidative stress and gut-derived lipopolysaccharides in children affected by paediatric autoimmune neuropsychiatric disorders associated with streptococcal infections

    DOI: 10.1186/s12887-020-02026-8

    PMID: 32188439

    		 Sample: Serum
  33. JOURNAL OF PAEDIATRICS AND CHILD HEALTH (2022) IF: 1.954
    Zonulin and claudin-5 levels in multisystem inflammatory syndrome and SARS-CoV-2 infection in children

    DOI: 10.1111/jpc.16033
    Sample: serum
  34. JGH Open (2021)
    Intestinal fatty acid-binding protein is a biomarker for diagnosis of biliary tract infection

    DOI: 10.1002/jgh3.12644
    Sample: blood
  35. Turkish Journal of Biochemistry-Turk Biyokimya Dergisi (2021) IF: 0.401
    The role of zonulin in the pathogenesis of diabetic retinopathy

    DOI: 10.1515/tjb-2020-0575
    Sample: serum
  36. JOURNAL OF OBSTETRICS AND GYNAECOLOGY (2020) IF: 0.8070
    Association of adverse outcomes of intrahepatic cholestasis of pregnancy with zonulin levels

    DOI: 10.1080/01443615.2020.1820463

    PMID: 33228404

    		 Sample: serum
  37. African Health Sciences (2021) IF: 0.927
    Gut permeability is associated with hypertension and measures of obesity but not with Endothelial Dysfunction in South African youth

    DOI: 10.4314/ahs.v21i3.26
    Sample: plasma
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Reviews/Q&A

  • Show all
  • Reviews
  • Q&A
Q

A************iSubmitted [ Jan 25 2024 ]

Asked: Hello, can I use this test on stool samples?

Reply

A

adminSubmitted [ Jan 25 2024 ]

Answered: Yes, this ELISA kit can be used to test feces samples.

Q

H**********aSubmitted [ Dec 13 2023 ]

Asked: Hii, i want to know the range of zonulin levels that we will consider as normal and above which we will consider that the zonulin levels are elevated. Thank you.

Reply

A

adminSubmitted [ Dec 13 2023 ]

Answered: For E-EL-H5560 kit, the reference value of normal human serum samples is 40-115ng/ml, according to our internal verification result. However, considering about the individual differences, we recommend that you set up your own normal sample group as control in order to determine whether the zonulin levels are elevated.

Q

S*eSubmitted [ Sep 13 2022 ]

Asked: In your previous answers to other questions, you said, \"the immunogen sequences of the capture antibody and detection antibody were human haptoglobin 19-406aa (#P00738).\" So that would mean that haptoglobin could produce a false positive or falsely elevated \"zonulin\" result in your test. How do you know you are detecting zonulin/pre-HB2 and not haptoglobin? You also said, \"The antigen is full-length recombinant pre-HP2. The expression host of the antigen is HEK 293 cell.\" Which antigen are you referring to in that second statement, since you already stated that the antigen which the capture antibody is specific for is haptoglobin rather than zonulin/pre-HP2? If the capture and detection antibodies react with both haptoglobin and pre-HP2, how is your assay specific for only pre-HP2? Thank you.

Reply

A

adminSubmitted [ Sep 13 2022 ]

Answered: We applied anti-ZOT antibodies in this kit, ZOT is an enterotoxin elaborated by Vibrio cholera, a zonulin analogue. It has been reported that this antibody specifically recognizes the human zonulin (pre-HP2) instead of the mature Haptoglobin. In addition, we verified that the kit does not have significant cross-reactivity with mature HP protein.

Q

p***aSubmitted [ Mar 22 2022 ]

Asked: Good morning, We were thinking of purchasing this kit for an internal study regarding intestinal permeability. We were wondering if you could provide with information regarding the validity of this kit and the accuracy in detecting the pre-haptoglobin-2. Would appreciate quick reply! Thank you for your time and help!

Reply

A

adminSubmitted [ Mar 22 2022 ]

Answered: The zonulin protein is the inactive precursor for HP2 and it is an uncleaved single-chain form. For our kit, it can detect the Precursor of haptoglobin 2, the immunogen sequences of the capture antibody and detection antibody were human haptoglobin 19-406aa (#P00738). The sensitivity report will be shared via email.

Q

h****nSubmitted [ Mar 15 2022 ]

Asked: Hi, I am wondering about the information on the variability of zonulin assay? And what are zonulin and occludin coefficients of variation? of other indexes of test-retest reliability?

Reply

A

adminSubmitted [ Mar 15 2022 ]

Answered: Hi Hicran, thanks for your message. For your question, not only zonulin or occludin kits, but all of our kits need to strictly check the precision evaluation parameters before shipment, that is, the intra-assay and inter-assay coefficients of variation of the kits are both <10%. Please refer to the manuals to obtain the coefficient of variation information of each kit. Thank you.

Q

T***mSubmitted [ Dec 10 2021 ]

Asked: Does your kit not recognize complement C3, as it was said, \"Top matches of the assays\' detected antigen included haptoglobin and complement C3 for the assay manufactured by CUSABIO (Wuhan, China) and complement C3 for the assay manufactured by Immundiagnostik AG (Bensheim, Germany) after the nature of the assays\' detected antigen was investigated using immunoprecipitation followed by mass spectrometric analysis and sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by protein staining. (Ajamian et al., PloS One, 2019)

Reply

A

adminSubmitted [ Dec 10 2021 ]

Answered: Thanks for your message. E-EL-H5560 Human Zonulin ELISA Kit Yes, the kit can recognize zonulin, but cannot recognize complement C3. We also found the document you mentioned in the attachment, these words don't mean one kit can recognize two proteins (zonulin, complement C3), which means the CUSABIO kit can detect serum zonulin levels, and our understanding literature says is human complement C3c, HP1-1, HP2-2 and recombinant zonulin (zon) standards were separated by SDS-PAGE and stained with Coomassie gel stain . If you want to test for C3, we recommend E-EL-H6054 Human C3(Complement Component 3) ELISA Kit for you. https://www.elabscience.com/p-human_c3_complement_component_3_elisa_kit-356179.html

Q

M*************iSubmitted [ Sep 09 2021 ]

Asked: Sorry for disturbing you again, I inquired few minutes ago about the reactivity of antibody to zonulin-recombinant. The another question is about What is the antigen when you make anti-zonulin antibody. I would appreciate your immediate attention to this matter.

Reply

A

adminSubmitted [ Sep 09 2021 ]

Answered: Dear Hayashi san,thanks for your message. This kit is specifically designed to detect Zonulin. The kit can be used to detect natural Zonulin. In theory, the kit also can be used for recombinant Zonulin. However, due to the influence of the immunogen sequence and the expression host of the recombinant protein, some recombinant Zonulin may not match the antibodies in the kit. Therefore, a pre-test is recommended. The antigen is full-length recombinant pre-HP2. The expression host of the antigen is HEK 293 cell. Best Regards, Y

Q

H*************oSubmitted [ Sep 09 2021 ]

Asked: Recently some ELISA kits for zonulin are in controversy because they actually don\'t detect zonulin. I would like to know that you confirm antibodies used in your kits capture zonulin-recombinants because I would like to purchase reliable products. Thank you.

Reply

A

adminSubmitted [ Sep 09 2021 ]

Answered: Dear Hayashi san,thanks for your message. We have replied to you in your second message column, please check it out, thank you! Best regards,

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