Last update: 3/20/12 IK
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TP104 – IF Troubleshooting
In order for us to help you better and faster, please provide us with as much information as
possible by filling out this form and returning to Abbiotec Technical Support Department by
email to techsupport@abbiotec.com or fax to 1 (858) 586-6252. Attach any relevant
experimental data (positive or negative) with legend.
1.
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General information
Customer name:
Name of the reagent:
Catalog Number:
Lot Number:
Date received:
Storage temperature:
Was the product aliquoted?
2.
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Specimen information
Species:
Organ/Tissue:
Primary cells:
Cell line:
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Fixation information
Immersion, perfusion, post-fixed, unfixed:
Fixative used:
Fixation time:
Fixation temperature:
4.
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Sectioning information
Cryoprotection:
Sections are mounted or free-floating?
Paraffin-embedded:
Fixed cryosections:
Fresh frozen:
Section thickness:
Please describe how the unstained samples were stored (time, temperature, storage
buffer, etc.,):
5. Antigen retrieval information
 Retrieval method:
Abbiotec, LLC • 7985 Dunbrook Rd, Ste A, San Diego, CA 92126 • USA
Ph: 858.586.0500 • Fax: 858.586.6252 • Email: info@abbiotec.com • WWW.ABBIOTEC.COM
Last update: 3/20/12 IK
Document1
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Retrieval buffer:
Retrieval time:
Retrieval temperature:
6.
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Permeabilization information
Permeablization method:
Permeabilization reagent:
Concentration:
Permeabilization time:
7.
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Endogenous biotin blocking information
Blocking reagent:
Concentration:
Blocking time:
8.
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Blocking before primary antibody information
Blocking reagent:
Blocking time:
Blocking temperature:
9.
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Primary antibody information
Dilution:
Diluent:
Was this a double/multiple immunofluroescent staining?
If yes, please describe all other primary antibodies used:
Length of incubation:
Temperature of incubation:
Washing after primary antibody incubation:
Washing buffer:
How many times was washed/how long:
10.
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Blocking before secondary antibody information
Blocking reagent:
Blocking time:
Blocking temperature:
11.
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Secondary antibody information
Supplier:
Secondary antibody species reactivity:
Secondary antibody is conjugated with:
Fluorochrome:
Abbiotec, LLC • 7985 Dunbrook Rd, Ste A, San Diego, CA 92126 • USA
Ph: 858.586.0500 • Fax: 858.586.6252 • Email: info@abbiotec.com • WWW.ABBIOTEC.COM
Last update: 3/20/12 IK
Document1
Page 3 of 4
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Dilution:
Diluent:
Was this a double/multiple immunofluroescent staining?
If yes, please describe all other secondary antibodies used:
Length of incubation:
Temperature of incubation:
Washing after secondary antibody incubation:
Washing buffer:
How many times was washed/how long:
12.
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Tertiary reagent information
Supplier:
Name of the reagent:
Tertiary reagent is conjugated with:
Fluorochrome:
Dilution:
Diluent:
Length of incubation:
Temperature of incubation:
Washing after tertiary reagent:
Washing buffer:
How many times was washed/how long:
13.
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Counterstaining information
Name of reagent:
Concentration:
Staining time:
14.
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Mounting/Coverslipping information
Mounting medium used for free-floating sections:
Coverslipping medium used:
Please describe, how the slides were stored between coverslipping and
photodocumentation (time, temperature, light exposure, etc.,):
15.
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Detection system information
Microscope/imaging system used:
Excitation source (lasers, mercury lamp, etc):
Filters used:
16. Results
 What is the expected localization of the signal in the tissue?
Abbiotec, LLC • 7985 Dunbrook Rd, Ste A, San Diego, CA 92126 • USA
Ph: 858.586.0500 • Fax: 858.586.6252 • Email: info@abbiotec.com • WWW.ABBIOTEC.COM
Last update: 3/20/12 IK
Document1
Page 4 of 4
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What is the expected subcellular localization of the signal?
Localization of the signal on the slide:
Background:
How many times this staining was tried?
Are the results the same every time?
Was this antibody previously working on these samples?
Was the secondary antibody working previously on other samples?
Positive and negative controls used:
Different dilutions tested for primary antibody:
Was a secondary antibody control (without primary antibody) run?
Abbiotec, LLC • 7985 Dunbrook Rd, Ste A, San Diego, CA 92126 • USA
Ph: 858.586.0500 • Fax: 858.586.6252 • Email: info@abbiotec.com • WWW.ABBIOTEC.COM