Anti-Cytokeratin 18 抗体 [C-04] (ab668)

Reviews (20)Q&A (9)References (65)

製品の概要

  • 製品名
    Anti-Cytokeratin 18 antibody [C-04]
    Cytokeratin 18 一次抗体 製品一覧
  • 製品の詳細
    Mouse monoclonal [C-04] to Cytokeratin 18
  • 由来種
    Mouse
  • 特異性
    Human Cytokeratin
  • アプリケーション
    適用あり: ICC, IHC-P, WB, Flow Cyt, IHC-Fr, IP, ICC/IFmore details
  • 種交差性
    交差種: Mouse, Rat, Sheep, Goat, Horse, Hamster, Cow, Cat, Dog, Human, Pig, Common marmoset
  • 免疫原

    Tissue/ cell preparation (Human). (Cytoskeleton preparation of epidermal carcinoma cell line A431).

  • ポジティブ･コントロール
    • A431 cells

製品の特性

アプリケーション

Our Abpromise guarantee covers the use of ab668 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

アプリケーション Abreviews 特記事項
ICC Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
WB Use at an assay dependent concentration. Predicted molecular weight: 45 kDa.
Flow Cyt Use 1µg for 106 cells.

Also see PMID 18946470.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
IHC-Fr 1/600. PubMed: 23769181
IP Use at an assay dependent concentration.
ICC/IF Use at an assay dependent concentration.

ターゲット情報

  • 機能
    Involved in the uptake of thrombin-antithrombin complexes by hepatic cells (By similarity). When phosphorylated, plays a role in filament reorganization. Involved in the delivery of mutated CFTR to the plasma membrane. Together with KRT8, is involved in interleukin-6 (IL-6)-mediated barrier protection.
  • 組織特異性
    Expressed in colon, placenta, liver and very weakly in exocervix. Increased expression observed in lymph nodes of breast carcinoma.
  • 関連疾患
    Defects in KRT18 are a cause of cirrhosis (CIRRH) [MIM:215600].
  • 配列類似性
    Belongs to the intermediate filament family.
  • 翻訳後修飾
    Phosphorylation at Ser-34 increases during mitosis. Hyperphosphorylated at Ser-53 in diseased cirrhosis liver. Phosphorylation increases by IL-6.
    Proteolytically cleaved by caspases during epithelial cell apoptosis. Cleavage occurs at Asp-238 by either caspase-3, caspase-6 or caspase-7.
    O-glycosylated at multiple sites; glycans consist of single N-acetylglucosamine residues.
  • 細胞内局在
    Cytoplasm > perinuclear region.
  • Information by UniProt
  • 参照データベース
    see all
  • 別名
    • Cell proliferation inducing gene 46 protein antibody
    • Cell proliferation inducing protein 46 antibody
    • Cell proliferation-inducing gene 46 protein antibody
    • CK 18 antibody
    • CK-18 antibody
    • CK18 antibody
    • CYK 18 antibody
    • CYK18 antibody
    • Cytokeratin 18 antibody
    • Cytokeratin endo B antibody
    • Cytokeratin-18 antibody
    • K 18 antibody
    • K18 antibody
    • K1C18_HUMAN antibody
    • KA18 antibody
    • Keratin 18 antibody
    • Keratin 18, type I antibody
    • Keratin D antibody
    • keratin, type I cytoskeletal 18 antibody
    • Keratin-18 antibody
    • Krt18 antibody
    see all

画像

  • Flow Cytometry - Anti-Cytokeratin 18 antibody [C-04] (ab668)
    Flow Cytometry - Anti-Cytokeratin 18 antibody [C-04] (ab668)
    Overlay histogram showing HCT116 cells stained with ab668 (red line). The cells were fixed with methanol (5 min) and then permeabilized with 0.1% PBS-Triton for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab668, 1µg/1x106 cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HCT116 cells fixed with 4% paraformaldehyde/permeabilized in 0.1% PBS-Triton used under the same conditions.

  • Immunocytochemistry/ Immunofluorescence - Anti-Cytokeratin 18 antibody [C-04] (ab668)
    Immunocytochemistry/ Immunofluorescence - Anti-Cytokeratin 18 antibody [C-04] (ab668)This image is courtesy of an anonymous Abreview

    ab668 staining Cytokeratin 18 in the human hepatocellular carcinoma cell line HepG2 by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with methanol/acetone (7:3) and blocked with 1% BSA for 1 hour at 20°C. Samples were incubated with primary antibody (1/100 in PBS ) for 1 hour at 37°C. An Alexa Fluor® 488-conjugated rabbit anti-mouse IgG polyclonal was used as the secondary antibody at 1/100 dilution.

    See Abreview

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cytokeratin 18 antibody [C-04] (ab668)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cytokeratin 18 antibody [C-04] (ab668)This image is courtesy of an Abreview submitted by Carl Hobbs

    ab668 staining Cytokeratin 18 in Cat lung tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 1% BSA for 10 minutes at 21°C; antigen retrieval was by heat mediation in citric acid. Samples were incubated with primary antibody (1/500 in TBS/BSA/azide) for 2 hours at 21°C. A Biotin-conjugated goat anti-mouse IgG polyclonal (1/200) was used as the secondary antibody.

    Coloured arrowheads indicate positivity.
    Good immunolabelling of bronchial tree epithelia (green), alveolar lining epithelium (red ).
    Goblet cells are negative (asterisk).
    Heavily stained structures are submucosal mucous glands.

    See Abreview

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cytokeratin 18 antibody [C-04] (ab668)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cytokeratin 18 antibody [C-04] (ab668)This image is courtesy of an Abreview submitted by Carl Hobbs

    ab668 staining cytokeratin 18 in Mouse epithelium tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 1% BSA for 10 minutes at 21°C; antigen retrieval was by heat mediation in a citrate buffer. Samples were incubated with primary antibody (1/500 in TBS/BSA/azide) for 2 hours at 21°C. A Biotin-conjugated Goat anti-mouse IgG polyclonal (1/200) was used as the secondary antibody.

    Composite image of d14 embryo shows developing epithelia.

    Upper image: positive simple epithelium of  renal tubules

    Lower image: negative stratified epithelium of epidermis

    See Abreview

  • Immunocytochemistry/ Immunofluorescence - Anti-Cytokeratin 18 antibody [C-04] (ab668)
    Immunocytochemistry/ Immunofluorescence - Anti-Cytokeratin 18 antibody [C-04] (ab668)Image courtesy of an anonymous Abreview.
    ab668 staining Cytokeratin 18 in horse primary bronchial epithelial cells by Immunocytochemistry/ Immunofluorescence. The cells were fixed in acetone and then blocked using 3% BSA for 10 hours at 4°C. Samples were then incubated with primary antibody at 1/100 for 1 hour at 22°C. The secondary antibody used was a goat anti-mouse conjugated to FITC used at a 1/200 dilution.

    See Abreview

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cytokeratin 18 antibody [C-04] (ab668)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Cytokeratin 18 antibody [C-04] (ab668)
    ab668 (2µg/ml) staining cytokeratin 18 in human skin using an automated system (DAKO Autostainer Plus). Using this protocol there is strong cytoplasmic staining of sweat coils.
    Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer citrate pH6.1 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako envision flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.

参考文献

This product has been referenced in:
  • Chua CW  et al. Differential requirements of androgen receptor in luminal progenitors during prostate regeneration and tumor initiation. Elife 7:N/A (2018). Read more (PubMed: 29334357) »
  • Shenoy PA  et al. The Somatostatin Receptor-4 Agonist J-2156 Alleviates Mechanical Hypersensitivity in a Rat Model of Breast Cancer Induced Bone Pain. Front Pharmacol 9:495 (2018). Read more (PubMed: 29867498) »
See all 65 Publications for this product

ab668 を使用した論文を発表された方は、こちらまでお知らせください。データシートに掲載させていただきます。

レビューと Q&A

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1-10 of 29 Abreviews or Q&A

Immunohistochemistry (Frozen sections) abreview for Anti-Cytokeratin 18 antibody [C-04]

 Excellent
Abreviews
abreview image
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Skin, oral epithelium)
Permeabilization
No
Specification
Skin, oral epithelium
Blocking step
Serum as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 0.25% · Temperature: 25°C
Fixative
Paraformaldehyde
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Abcam user community

Verified customer

投稿 Dec 29 2017

Western blot abreview for Anti-Cytokeratin 18 antibody [C-04]

 Good
Abreviews
Abcam guarantees this product to work in the species/application used in this Abreview.
abreview image
Application
Western blot
Sample
Human Cell lysate - whole cell (T24 cells)
Gel Running Conditions
Reduced Denaturing (10)
Loading amount
35 µg
Specification
T24 cells
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: RT°C
Read More

Abcam user community

Verified customer

投稿 Jun 12 2017

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-Cytokeratin 18 antibody [C-04]

 Poor
Abreviews
abreview image
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citric acid
Sample
Pig Tissue sections (Small intestine)
Specification
Small intestine
Permeabilization
No
Fixative
Formaldehyde
Read More

Mr. Carl Hobbs

Verified customer

投稿 Sep 11 2014

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-Cytokeratin 18 antibody [C-04]

 Excellent
Abreviews
abreview image
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citric acid
Sample
Cat Tissue sections (Lung)
Specification
Lung
Permeabilization
No
Fixative
Formaldehyde
Read More

Mr. Carl Hobbs

Verified customer

投稿 Jan 13 2014

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) abreview for Anti-Cytokeratin 18 antibody [C-04]

 Excellent
Abreviews
abreview image
Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Blocking step
BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citric acid
Sample
Goat Tissue sections (Lung)
Specification
Lung
Permeabilization
No
Fixative
Formaldehyde
Read More

Mr. Carl Hobbs

Verified customer

投稿 Jan 13 2014

Immunohistochemistry (PFA perfusion fixed frozen sections) abreview for Anti-Cytokeratin 18 antibody [C-04]

 Excellent
Abreviews
abreview image
Application
Immunohistochemistry (PFA perfusion fixed frozen sections)
Blocking step
BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 1:1000 · Temperature: RT°C
Sample
Rabbit Tissue sections (eye)
Specification
eye
Permeabilization
No
Fixative
Formaldehyde
Read More

Dr. Alex Zagariya

Verified customer

投稿 Jan 06 2014

Question


Thank you for your reply! I know Ab668 is a popular one, but most mouse
antibodies have background when stain on mouse tissue.
The Ab668 also has high background in our mouse olfactory epithelium
tissue. Please check the attachment file. That's why I want to order a
rabbit anti-ck18.
Our protocol as follows:
1. olfactory epithelium tissue fixed in 4%PFA at 4C overnight
wash and equilibrated in sucrose/PBS, embeded in OCT.
2. Cryosections were prepared at 10 um. 4%PFA postfixed for 5min.
3. Washed in PBS and blocked in 10% Normal Goat serum or Donkey serum plus
1%BSA. 4C overnight.
4. Ms-antiCK18(Ab668) 1:100 in blocking buffer 4C 48 hours
5. Washed in PBS 4x and incubated with Gt-anti Ms Alex fluo 546 Dilution
1000, incubated 1.5hour at Room temperature. Then wash in PBS 4X.
According to the publications, CK18 stain the cells on the apical layer in
Olfactory epithelium. But Ab668 stained everywhere.

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Answer

Hello! Please find below our mouse on mouse staining tips. This may help you to improve your results. Additionally I would recommend you reduce the primary incubation to overnight, and incubate the antibodies in a solution of TBS plus 1% BSA.

We cannot vouch for the Millipore version of the clone but we have tested ab32118 and found that it does not react in mouse, and have placed this information on our datasheet to inform our customers.



Tips for eliminating background when staining mouse tissue with a mouse monoclonal



View a pdf version of this page
Staining of mouse tissue using mouse antibody is a complicated process as high levels of background are often observed. It is notoriously difficult to eliminate this background.
Much of the background is caused by secondary antibody binding to endogenous mouse IgG in the tissue being stained, and to Fc receptors on B cells, plasma cells and macrophages.
Abcam is unable to guarantee monoclonal mouse antibodies on mouse tissue (unless stated on the datasheet). However, there are a few tips to try and reduce this background should mouse on mouse staining be necessary:

1. Blocking of endogenous IgG



Prepare tissue sections as usual.
At the usual blocking step, block with serum (from same species as the secondary antibody) for 30 min at room temperature.
Wash 3 X 2 min with PBS Tween 20.
Incubate sections with an unconjugated AffiniPure Fab fragment Anti-Mouse IgG (H+L) for 1 hr at room temperature, or overnight 4°C . Try this blocking antibody at 0.1 mg/ml although the optimal concentration will need to be assessed by the end user.Proceed with antibody staining.




2. Blocking endogenous Fc receptors
There are kits available for this which use F(ab) monomeric secondary antibodies.

Other tips that can be used to decrease general background:


Incubate sections with 1% Triton (in PBS) at room temperature for 30 min to ‘clean’ the tissue.
Use TBS –Tween 20 as a washing buffer. This often gives less background than using PBS-Tween 20.

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Question

1. The concentration of sheep serum used is 1:1000, diluted in PBS+1% TritonX. We block at room temperature for 90 minutes.

2. Antigen retrieval was performed at 96C for 25 minutes. We haven’t performed a time series, we just went off several methods which recommended 25 minutes for antigen retrieval. There was no difference between the two pHs. The pH 10 buffer we used was in fact a Tris based buffer, not a citrate based buffer.

3. We haven’t tested the secondary antibody with known primary antibodies which work as yet, as our lab has limited primary antibodies raised in mouse. We do however have a goat-anti-mouse fluorescent antibody which does work, and we will be trying this in the near future.



Thanks,

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Answer

Thank you for your reply.


I am sorry to hear that you have difficulties getting satisfactory results form this antibody.

The details you have kindly provided will enable us to investigate this case for you and this is also helpful in our records for monitoring of quality.

Reviewing this case, I would like to offer some suggestions to help optimize the results from ab668.

1.) I suggest to decrease the amount of Triton X used in the buffers. 0.025% Triton X-100 is used in standard protocols. Triton X is a quite harsh detergent.

2.) Secondary anti pan IgG antibodies sometimes bind some isotypes better than others. I therefore suggest to try a different secondary antibody and use an IgG1 specific antibody.

Please let me know how you are getting on.

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Question

Hello, I am attempting to use Ab668 - Cytokeratin 18 on rat liver sections, which were fixed in formalin for 24 hours and paraffin processed. After several troubleshoots, we are unable to get the antibody to work, even at 1:100. Primary antibody conditions are always diluted in PBS + 1% triton, overnight at 4C. We block in sheep serum (1:1000, diluted in PBS + 1% triton), have tried a pH 6 citrate buffer and pH 10 sodium citrate buffer, both at 96C for antigen retrieval, have used standard DAB detection (DAKO) and amplification via the Vector ABC kit. Secondary antibody used is a goat-anti-mouse HRP, at 1:1000, diluted in PBS for 1hour at room temp. All washes are conducted with PBS and we always have a no-primary antibody negative control. We also always block for biotin via 0.3% hydrogen peroxide in PBS, for 15 mins at room temp. This step has always been conducted before the secondary antibody incubation. Every time, there is no staining with this antibody, and sections incubated with primary antibody look like the no primary negative control. All signs point to the primary antibody not working. Do you have any suggestions on how to proceed?

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Answer

Thank you for your enquiry.

I am sorry to hear that this antibody is not providing satisfactory results.

The details provided will enable us to investigate this case and will provide us with vital information for monitoring product quality.

Having reviewed this case, I would appreciate if you can confirm some further details:

1. What is the sheep serum's concentration and for how long do you block?

2.How long wasthe retrieval? Did you perform a time series? Do you see any difference between the two pHs?

3. Do you know if the secondary antibody is working?

I thank you for your cooperation.

Read More

Question

Hi,

For Cytokeratin 18 antibody [C-04] (ab668), does it specifically reacts with human origin epithelial cells not murine origin cells ?

Thanks

Read More
Answer

Thnak you for your inquiry.

I have to confirm that ab668 recognizes both human and mouse Cytokeratin 18 as stated on the datasheet.

If you are looking for an antibody that only binds to human Cytokeratin 18, I cannot recommend ab668.

I can recommend to following antibodies that react with human Cytokeratin 18, but do not react with mouse:

ab133302

https://www.abcam.com/index.html?datasheet=133302 (or use the following: https://www.abcam.com/index.html?datasheet=133302).

ab133272

https://www.abcam.com/index.html?datasheet=133272 (or use the following: https://www.abcam.com/index.html?datasheet=133272).

ab133263

https://www.abcam.com/index.html?datasheet=133263 (or use the following: https://www.abcam.com/index.html?datasheet=133263).

I hope this information is helpful and wish you good luck with your research.

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1-10 of 29 Abreviews or Q&A

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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