Overview

Applications

Fast track antibodies constitute a diverse group of products that have been released to accelerate your research, but are not yet fully characterized. They have all been affinity purified and show high titre values against the immunizing peptide (by ELISA). Fast track terms of use

Application Abreviews Notes
WB 1/1000. Detects a band of approximately 55 kDa (predicted molecular weight: 58 kDa).

Target

  • Function

    Catalyzes a rate-limiting step in cholesterol catabolism and bile acid biosynthesis by introducing a hydrophilic moiety at position 7 of cholesterol. Important for cholesterol homeostasis.
  • Tissue specificity

    Detected in liver.
  • Pathway

    Lipid metabolism; bile acid biosynthesis.
  • Sequence similarities

    Belongs to the cytochrome P450 family.
  • Cellular localization

    Endoplasmic reticulum membrane. Microsome membrane.
  • Information by UniProt
  • Database links

  • Alternative names

    • Cholesterol 7 alpha hydroxylase antibody
    • Cholesterol 7 alpha monooxygenase antibody
    • Cholesterol 7-alpha-hydroxylase antibody
    • Cholesterol 7-alpha-monooxygenase antibody
    • CP7A antibody
    • CP7A1_HUMAN antibody
    • CYP 7 antibody
    • CYP7 antibody
    • CYP7A1 antibody
    • CYPVII antibody
    • Cytochrome P450 7A1 antibody
    • Cytochrome P450, family 7, subfamily A, polypeptide 1 antibody
    • Cytochrome P450, subfamily VIIA (cholesterol 7 alpha monooxygenase), antibody
    • Cytochrome P450, subfamily VIIA (cholesterol 7 alpha-monooxygenase), polypeptide 1 antibody
    • MGC126826 antibody
    • MGC138389 antibody
    see all

Images

This Fast-Track antibody is not yet fully characterised. These images represent inconclusive preliminary data.

  • All lanes : Anti-CYP7A1 antibody (ab65596) at 1/1000 dilution

    All lanes : Mouse liver lysate

    Lysates/proteins at 30 µg per lane.

    Secondary
    All lanes : HRP labeled goat anti-rabbit at 1/5000 dilution

    Developed using the ECL technique.

    Performed under non-reducing conditions.

    Predicted band size: 58 kDa
    Additional bands at: 40 kDa (possible non-specific binding), 70 kDa (possible non-specific binding)



    Blocking: 5% Milk

    See Abreview

References

This product has been referenced in:

  • Yetti H  et al. Bile acid detoxifying enzymes limit susceptibility to liver fibrosis in female SHRSP5/Dmcr rats fed with a high-fat-cholesterol diet. PLoS One 13:e0192863 (2018). WB ; Rat . Read more (PubMed: 29438418) »
  • Niu S  et al. Beta-Arrestin 1 Mediates Liver Thyrotropin Regulation of Cholesterol Conversion Metabolism via the Akt-Dependent Pathway. Int J Endocrinol 2018:4371396 (2018). Read more (PubMed: 29853881) »
See all 16 Publications for this product

Customer reviews and Q&As

1-5 of 5 Abreviews or Q&A

Application
Western blot
Sample
Mouse Tissue lysate - whole (liver)
Gel Running Conditions
Non-reduced Denaturing (12%)
Loading amount
30 µg
Specification
liver
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 37°C

Abcam user community

Verified customer

Submitted Jul 05 2017

Application
Western blot
Loading amount
50 µg
Gel Running Conditions
Reduced Denaturing (7.5)
Sample
Rat Tissue lysate - whole (liver)
Specification
liver
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 7.5%

Abcam user community

Verified customer

Submitted Sep 09 2013

Answer



ab65596 es un llamado “Fast Track” antibody. Se trata de anticuerpos que no se han caracterizado completamente, y que se han añadido al catálogo antes de realizar la validación completa por la necesidad urgente de añadir anticuerpos contra la proteína concreta.

Gracias a que no están completamente caracterizados su precio es reducido y no están cubiertos por la garantía Abpromise. Solo se han testado en el laboratorio en ELISA, y cualquier publicación o referencia de los usuarios del mismo se compensa con Abpoints extra canjeables.

Los términos y condiciones de estos anticuerpos los podéis consultar en su datasheet.

Por tanto, en respuesta a tu pregunta, este anticuerpo no se ha usado en Western Blot, ni con tejidos de ratón, y tampoco podemos ofrecer códigos de descuento Abtrial. Sin embargo, si estáis interesados en usarlo en esta especie / aplicación, y enviarnos el Abreview, sí que a cambio os beneficiareis de 950 Abpoints en vuestra cuenta. El anticuerpo está actualmente en stock.

En cualquier caso, hay disponibles otros dos anticuerpos que si se han validado en WB con lisados de ratón: ab106060 y ab65596.

Read More

Question

Dear Technical support,

I hope this email finds you well.

We've received a product complaint which we are hoping that you will be able to help address. Attached is the completed questionnaire.

With many thanks and best regards,



1) Abcam product code: ab65596



2) Abcam order reference number or product batch number: 831470



3) Description of the problem: There seems to be bands in the beginning. After that, the WB film either turns black entirely or no band shows up.



4) Sample preparation

Type of sample: whole cell lysates

Lysis buffer: RIPA

Protease inhibitors: Halt(tm)Protease & PhosphataseInhibitor Cocktail

Phosphatase inhibitors: Halt(tm)Protease & PhosphataseInhibitor Cocktail

Reducing agent: Loading buffer (Bio-Rad) & β-mercaptoethanol

Boiling for ≥5 min? Yes, 95˚C for 15 minutes

Protein loaded: 50ug / lane

Positive control: HepG2, rat liver

Negative control: Blank lane



5) Percentage of gel: 8% separating. 5% stacking.

Type of membrane: PVDF

Protein transfer verified : Yes, using ladder as a gauge

Blocking agent and concentration: 5% milk in 1X TBS w/ 0.05% Tween 20

Blocking time: 1 hour

Blocking temperature: Room temperature



6) Primary antibody

Concentration or dilution: 1:1000, 1:1500, 1:2000

Diluent buffer: 1X TBS w/ 0.05% Tween 20

Incubation time: Overnight

Incubation temperature: 4˚C




7) Secondary antibody

Species: Goat

Reacts against: anti-rabbit

Concentration or dilution: 1:10000, 1:15000, 1:20000

Diluent buffer: 1X TBS w/ 0.05% Tween 20

Incubation time: 1 hour

Incubation temperature: Room temperature

Fluorochrome or enzyme conjugate: Enzyme conjugate



8) Washing after primary and secondary antibodies:

Buffer: twice with 1X TBS w/ 0.05% Tween 20, thrice with 1X TBS

Number of washes: 5, 5 minutes each wash



9) Detection method: ECL



10) How many times have you run this staining? Seven

Do you obtain the same results every time? No, in the beginning got bands, after that, WB film either turns black entirely or no bands at all.

What steps have you altered to try and optimize the use of this antibody?


Primary antibody

Secondary antibody

Blocking


Document attachment: Attaching images of your blot is strongly recommended and can greatly speed up our investigation of your problem.

I have attached film picture when there were bands (or so I think, that they should be the correct bands). I did not attach those that are entirely black or entirely blank (w/o any bands).

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Answer

Thank you for taking the time to complete our questionnaire and contact us. I am sorry to hearthe customer hashad difficulty obtaining satisfactory results from 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 ab65596. I would also appreciate if you can confirm some further details:

1. I am sorry to confirm that this is a FastTrack antibody, not yet tested and guaranteed for rat or western blot. As our FastTrack antibodies have not been fully characterized at Abcam, we cannot guarantee that the antibody will work in any application other than ELISA against the immunizing peptide and I am sorry they are therefore not covered by the full Abpromise guarantee. Therefore, we are regrettably not able to provide a credit or refund on this occasion.

As an alternative, the customer may like to consider submitting an Abreview, for which they can obtain 900 Abpoints for data ona Fast Track antibody.

2. The secondary antibody mentioned on the form is stated as an anti-mouse. Could you confirm if this is correct? Please provide the supplier and catalog code for the secondary antibody.

As ab65596 is a rabbit polyclonal you will need to use an anti-rabbit IgG secondary antibody. An anti-mouse secondary will not work for detection of the rabbit polyclonal primary.

3. Is the secondary antibody working well with other primary antibodies? If you are getting background from a correct secondary, I can suggest it would be important to include a no primary control to assess if the background is from non specific binding of the secondary. The concentration of secondary may need to be reduced in order to optimize the results.

I hope this information is helpful, thank you for your cooperation. Should the suggestions not improve the results, please do not hesitate to contact me again with the further requested details.

Read More
Application
Western blot
Sample
Mouse Tissue lysate - whole (liver)
Loading amount
30 µg
Specification
liver
Gel Running Conditions
Reduced Denaturing (4-20% gel)
Blocking step
Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C

Abcam user community

Verified customer

Submitted Jan 19 2011

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