Overview

  • Product name
    Anti-Islet 1 antibody - Neural Stem Cell Marker
    See all Islet 1 primary antibodies
  • Description
    Rabbit polyclonal to Islet 1 - Neural Stem Cell Marker
  • Host species
    Rabbit
  • Specificity
    ab20670 might also detect rat and human Islet 2 protein as the immunogen used to raise this ab20670 is 86% identical to rat and human Islet 2. This has not been tested.
  • Tested applications
    Suitable for: IHC-FoFr, IHC-P, ICC/IF, IHC-Frmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human, Apteronotus leptorhynchus
    Predicted to work with: Chicken, Zebrafish
  • Immunogen

    Synthetic peptide conjugated to KLH derived from within residues 300 to the C-terminus of Human Islet 1.

    Read Abcam's proprietary immunogen policy (Peptide available as ab21996.)

  • Positive control
    • IHC-P: FFPE mouse embryo E12.

Properties

Applications

Our Abpromise guarantee covers the use of ab20670 in the following tested applications.

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

Application Abreviews Notes
IHC-FoFr Use at an assay dependent concentration. PubMed: 20096094
IHC-P Use a concentration of 0.5 - 1 µg/ml. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
ICC/IF Use a concentration of 2 µg/ml.
IHC-Fr 1/500.

Target

Images

  • IHC image of Islet 1 staining in a section of formalin fixed, paraffin embedded mouse embryo E12, performed on a Leica Bond™ system using the standard protocol B. The section was pre-treated using heat mediated antigen retrieval (EDTA based pH 9.0 solution, epitope retrieval solution 2) for 20 mins. The section was then incubated with ab20670, 0.5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

     

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • IHC image of Islet 1 staining in a section of formalin fixed, paraffin embedded mouse embryo E12, performed on a Leica Bond™ system using the standard protocol B. The section was pre-treated using heat mediated antigen retrieval (EDTA based pH 9.0 solution, epitope retrieval solution 2) for 20 mins. The section was then incubated with ab20670, 0.5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

     

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • ab20670 2µg/ml staining ISLET1 in human pancreas using an automated system (DAKO Autostainer Plus). Using this protocol there is strong nuclear and weak cytoplasmic staining primarily in the pancreatic islet.
    Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer EDTA pH 9.0 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.
  • Dorsal root ganglion explants were dissected from 16 day-old rat embryos and cultured for 6 hours in vitro with Neurobasal Medium containing B27 supplement.     

    Nuclei stained positive for anti-Islet 1 antibody ab20670 at 2µg/ml. As would be expected, not all cells in this preparation were Islet 1-positive. Pre-incubation of ab20670 with the immunizing peptide ab21996 resulted in complete blocking of the antibody.

    Green = ab20670
    Blue = To-pro-3 nuclear stain

    The level of magnification is different in each image.    

  • Immunocytochemistry/ Immunofluorescence analysis of adult mouse cardiac fibroblasts labeling Islet 1 with ab20670 at 1/100 dilution (green). Samples were fixed using 4% PFA with 0.25% TritonX-100. Scale bar is 100 µM.

  • Immunohistochemistry (Frozen sections) analysis of mouse brain tissue sections labeling Islet 1 with ab20670 at 1/200 dilution. The tissue was fixed with paraformaldehyde followed by blocking with 5% serum for 1 hour at 25°C. The tissue was incubated with ab20670 in PBST for 12 hours at 4°C. A polyclonal donkey anti-rabbit Alexa Fluor® 594 secondary antibody was used at 1/1000 dilution.

    See Abreview

  • Immunocytochemistry/ Immunofluorescence analysis of mouse dorsal root ganglia (DRG) neurons labeling Islet 1 with ab20670 at 1/200 dilution. Cells were fixed with paraformaldehyde and permeabilized with Tween 20. Blocking of the cells was performed with 2% serum for 30 minutes at 21°C, followed by incubation with ab20670 for 18 hours at 4°C. A polyclonal goat anti-rabbit Alexa Fluor® 568 secondary antibody was used at 1/1000 dilution. DAPI was used to counterstain.

    See Abreview

  • Immunohistochemical analysis of paraffin-embedded mouse E11.5 somites labelling Islet 1 with ab20670 at 1/100 dilution, followed by BioGenex polymer-HRP reagent (undiluted according to manufacturer).

  • ab20670 detected Islet 1 in the nucleus of frozen sections of E11.5 mouse embryonic brain using 1 ug/ml of antibody. Brighter staining may be achieved using a greater concentration of antibody, e.g. 2.5 or 5 ug/ml.

References

This product has been referenced in:
  • Alshawaf AJ  et al. Phenotypic and Functional Characterization of Peripheral Sensory Neurons derived from Human Embryonic Stem Cells. Sci Rep 8:603 (2018). Read more (PubMed: 29330377) »
  • Hsu F  et al. Rab5 and Alsin regulate stress-activated cytoprotective signaling on mitochondria. Elife 7:N/A (2018). Read more (PubMed: 29469808) »
See all 35 Publications for this product

Customer reviews and Q&As

1-10 of 22 Abreviews or Q&A

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry
Sample
Mouse Cell (Differentiated mES cell)
Permeabilization
Yes - 0.2% Triton X
Specification
Differentiated mES cell
Blocking step
BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 25°C
Fixative
Formaldehyde

Abcam user community

Verified customer

Submitted Nov 23 2016

Application
Immunocytochemistry/ Immunofluorescence
Blocking step
Serum as blocking agent for 15 minute(s) · Concentration: 5% · Temperature: RT°C
Sample
Human Cell (Human pluripotent stem cell derived cardiomyocyte)
Specification
Human pluripotent stem cell derived cardiomyocyte
Permeabilization
Yes - saponin
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Jan 12 2015

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Antigen retrieval step
Heat mediated
Sample
Spotted Catshark (Scyliorhinus canicula) Tissue sections (brain cells)
Specification
brain cells
Permeabilization
No
Fixative
Paraformaldehyde

Dr. Liam Rasch

Verified customer

Submitted Nov 05 2013

Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (Dorsal Root Ganglia (DRG) neurons)
Specification
Dorsal Root Ganglia (DRG) neurons
Fixative
Paraformaldehyde
Permeabilization
Yes - tween 20
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 2% · Temperature: 21°C

Abcam user community

Verified customer

Submitted Oct 18 2012

Application
Western blot
Sample
Apteronotus leptorhynchus Tissue lysate - whole (Brain)
Loading amount
50 µg
Specification
Brain
Gel Running Conditions
Reduced Denaturing (4-15%)
Blocking step
Milk as blocking agent for 14 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C

Abcam user community

Verified customer

Submitted Sep 28 2012

Answer

Thank you for your message.

I am pleased the results have improved. In order to help further, I would appreciate if you are able to send an attached image which will help me to assess the results.

Thank you for your continued cooperation. I look forward to hearing from you.

Read More

Question

Hi,
I hope this helps! Cheers

Order Details
Antibody code: ab20670
Lot number: GR82364-1
General Information
Antibody storage conditions (temperature/reconstitution etc)
The antibody was aliquoted and stored at -20C. The work I have been
doing used a fresh aliquot.
Description of the problem (high background, low signal, non-specific
satining etc.)
Non-specific staining and very high background
Sample (Species/Tissue/Cell Type/Cell Line etc.)
Human embryonic stem cell derived neurons
Fixation of sample (Ethanol/Methanol/Acetone/Paraformaldehyde/Other/
Duration etc.)
4% paraformaldehyde
How long for? We recommend 10 minutes only. 15minutes
Antigen retrieval (Enzymatic method, Heat mediated technique etc.) None
Permeabilization step
0.1% tritonX in PBS
How long for? 15 minutes
Blocking conditions (Buffer/time period, Blocking agent etc.)
10% goat serum for an hour
Primary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation
time, Wash step)
Overnight at 4C
1% goat serum in PBS/TritonX at a 1:400 concentration
Have you tried different concentrations? I haven't tried lowering the
concentration yet- this was going to be my next option.
Secondary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation
time, Wash step)
2 hours at room temperature. alexafluor goat anti-rabbit 488 at 1:500.
Is the secondary antibody working well with other primary antibodies?
Yes
Detection method : microscope
Positive and negative controls used (please specify) Negative control
was without primary antibody and it was clean. Also all the cells in
my culture were positive and this shouldn't be the case
Optimization attempts (problem solving)
How many times have you tried the IHC? Once
Have you run a "No Primary" control?
Yes
Do you obtain the same results every time?
Yes
What steps have you altered?
Additional Notes

Read More
Answer

Thank you for taking the time to complete our questionnaire and provide further information.

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 optimise the results. I would also appreciate if you can confirm some further details:

1. I would appreciate if you are able to provide an image which would help me to assess the results.

2. Could you confirm the wash steps? We recommend to wash 3 or 4 times at each appropriate wash step using PBS containing 0.2% Tween. This will help to wash away any excess antibody and also keep the cells permeabilized.

3. I agree that reducing antibody concentration would be beneficial to try. Try 1:1000.

4. Try reducing the secondary antibody concentration aswell.

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 information, and details of how you would like to proceed.

Read More

Answer

Thank you for taking the time to contact us. I am sorry to hear you have had difficulty obtaining satisfactory results from this antibody.

I appreciate the time you have spent in the laboratory and understand your concerns. It is regrettable the results have not been succesful.

I would like to reassure you that we monitor feedback closely on a weekly basis and we are not currently concerned about the general quality of this antibody or this batch.

I would also like to reassure you that this antibody is tested and covered by our guarantee for ICC-IF and human samples. Before deciding how to proceed, I would like to investigate this particular case further for you, and also obtain some further information for our quality records.

In order to do this, I have enclosed a questionnaire below. I would appreciate if you could complete this. It will help you put the information we require together very easily. I have completed the sections for which information has already been provided.

I would appreciate if you are also able to provide an image which will help us to assess the results.

In the event that a product is not functioning in the tested applications and species cited on the product data sheet (and the problem has been reported within 6 months of purchase), we will be pleased to provide a credit note, free of charge replacement or refund.

Thank you for your time and cooperation. We look forward to receiving the completed questionaire.

Order Details
Antibody code: ab20670
Lot number: GR82364-1
Purchase order number
or preferably Abcam order number:
General Information
Antibody storage conditions (temperature/reconstitution etc)
Description of the problem (high background, low signal, non-specific satining etc.)
Sample (Species/Tissue/Cell Type/Cell Line etc.)
Fixation of sample (Ethanol/Methanol/Acetone/Paraformaldehyde/Other/Duration etc.)
4% paraformaldehyde
How long for? We recommend 10 minutes only.
Antigen retrieval (Enzymatic method, Heat mediated technique etc.)
Permeabilization step
0.1% tritonX in PBS
How long for? We would recommend to permeabilize for 10 minutes only. Also, to include a more gentle detergent, such as 0.2% Tween in the wash buffer and antibody dilution buffer. this will help to keep the antibody solubilised and the cells permeabilised. Wash 3 times for 5 minutes at each appropriate wash step.
Blocking conditions (Buffer/time period, Blocking agent etc.)
10% goat serum for an hour
Primary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)
Overnight at 4C
1% goat serum in PBS/TritonX at a 1:400 concentration
Have you tried different concentrations?
Secondary Antibody (Manufacturer/Species/Diluent/Dilution/Incubation time, Wash step)
2 hours at room temperature. alexafluor goat anti-rabbit 488 at 1:500.
Is the secondary antibody working well with other primary antibodies?
Detection method
Positive and negative controls used (please specify)
Optimization attempts (problem solving)
How many times have you tried the IHC?
Have you run a "No Primary" control?
Yes No
Do you obtain the same results every time?
Yes No
What steps have you altered?
Additional Notes
We would appreciate if you are also able to provide and image which woudl help us to assess the results

Read More

Answer

Thank you for contacting Abcam.

Please find attached the Certificate of Compliance for ab20670, lot #*.

If there is anything else I can help you with, please let me know.

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Tissue sections (Adult heart)
Specification
Adult heart
Fixative
Formaldehyde
Permeabilization
Yes - 0.5% Tx100 in PBS
Blocking step
Serum as blocking agent for 1 hour(s) and 30 minute(s) · Concentration: 1% · Temperature: 20°C

Abcam user community

Verified customer

Submitted Jun 19 2012

1-10 of 22 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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