ab Universal HIER antigen retrieval reagent (10X)

Transcription

1 ab Universal HIER antigen retrieval reagent (10X) Instructions for use: For heat-induced epitope retrieval of formalin-fixed paraffin-embedded tissues This product is for research use only and is not intended for diagnostic use. Last Updated 24 September 2015

2 Table of Contents INTRODUCTION 1 1. BACKGROUND 1 GENERAL INFORMATION 2 2. PRECAUTIONS 2 3. STORAGE AND STABILITY 2 4. LIMITATIONS 3 5. MATERIALS SUPPLIED 3 6. MATERIALS REQUIRED, NOT SUPPLIED 4 7. TECHNICAL HINTS 5 ASSAY PROCEDURE 6 8. ASSAY PROCEDURE 6 DATA ANALYSIS 7 9. NOTES 7

3 INTRODUCTION INTRODUCTION 1. BACKGROUND Abcam s Universal HIER antigen retrieval reagent (10X) is a heatinduced epitope retrieval (HIER) solution that is compatible with virtually all antibodies and eliminates the need for multiple buffers, including citrate buffer, EDTA or high ph Tris buffers. Antibody titers are doubled or tripled when compared to citrate buffer, ph 6.0. Universal HIER antigen retrieval reagent (10X) incorporates a color-coded high temperature ph indicator solution, allowing the end-user to check that the solution is at the correct dilution and ph by visual inspection. This product is specially formulated for superior ph stability at high temperatures in order to help prevent the possibility of losing phsensitive antigens. Universal HIER antigen retrieval reagent (10X) is non-toxic, non-flammable, odorless and sodium azide- and thimerosalfree. The Universal HIER antigen retrieval reagent (10X) is a buffered solution that is intended for laboratory use in heat induced antigen retrieval of formalin-fixed paraffin-embedded (FFPE) tissues for immunohistochemistry (IHC) procedures. The clinical interpretation of any staining or its absence should be complimented by morphological studies using proper controls and should be evaluated within the context of the patient's clinical history and other diagnostic tests by a qualified pathologist. ab Universal HIER antigen retrieval reagent (10X) 1

4 GENERAL INFORMATION GENERAL INFORMATION 2. PRECAUTIONS Please read these instructions carefully prior to beginning the assay. We understand that, occasionally, experimental protocols might need to be modified to meet unique experimental circumstances. However, we cannot guarantee the performance of the product outside the conditions detailed in this protocol booklet. Please review the Safety Datasheet (SDS) provided with the product for information on how to handle and dispose of this reagent. Observe good laboratory practices. Gloves, lab coat, and protective eyewear should always be worn. Never pipet by mouth. Do not eat, drink or smoke in the laboratory areas. All biological materials should be treated as potentially hazardous and handled as such. They should be disposed of in accordance with established safety procedures. Avoid contacting the skin and mucous membranes with reagents and specimens. If reagents or specimens come in contact with sensitive areas, wash with copious amounts of water. 3. STORAGE AND STABILITY Store product at room temperature. See product label for expiry date. ab Universal HIER antigen retrieval reagent (10X) 2

5 GENERAL INFORMATION 4. LIMITATIONS Product intended for research use only. Not for use in diagnostic procedures. Do not mix reagents or materials from other product lots or vendors. The protocols for a specific application can vary. These include, but are not limited to: fixation, heat-retrieval method, incubation times, tissue section thickness and detection kit used. 5. MATERIALS SUPPLIED Item Amount Storage Condition Universal HIER antigen retrieval reagent (10X) 100 ml RT ab Universal HIER antigen retrieval reagent (10X) 3

6 GENERAL INFORMATION 6. MATERIALS REQUIRED, NOT SUPPLIED These materials will be required to successfully perform this assay: Deionised water Pipettes and pipette tips, including multi-channel pipette Assorted glassware for the preparation of reagents and buffer solutions Microscope slides, positively charged Drying oven Positive and negative tissue controls Xylene (or xylene substitute) Ethanol or reagent alcohol Pressure cooker Deionized or distilled water Wash buffer (TBS/PBS) Enzymes for digestion Avidin-Biotin Blocking Kit (if using Labeled Streptavidin-Biotin (LSAB) or avidin-biotin complex kits for detection) Peroxidase block Protein block or suitable serum Primary antibody Negative control reagents Detection kits Chromogens Hematoxylin Bluing reagent Mounting medium ab Universal HIER antigen retrieval reagent (10X) 4

7 GENERAL INFORMATION 7. TECHNICAL HINTS Review the protocol completely to confirm this product meets your requirements. Please contact our Technical Support staff with any questions. Avoid foaming or bubbles when mixing or reconstituting components. Avoid cross contamination of samples or reagents by changing tips between sample, standard and reagent additions. Ensure all reagents and solutions are at the appropriate temperature before starting the assay. Make sure all necessary equipment is switched on and set at the appropriate temperature. ab Universal HIER antigen retrieval reagent (10X) 5

8 ASSAY PROCEDURE ASSAY PROCEDURE 8. ASSAY PROCEDURE Equilibrate all materials and prepared reagents to correct temperature prior to use Deparaffinize tissues and hydrate with water. If necessary, block for endogenous peroxidase and wash in deionised water Dilute concentrated Universal HIER antigen retrieval reagent (10X) at a ratio of 1:10 (1 ml Universal HIER antigen retrieval reagent (10X) to 9 ml of deionized water) Place slides into 1X retrieval solution in a slide container (e.g. Coplin Jar, staining dish or metal slide canister) Retrieve sections using heat. Follow the recommendations on the antibody data sheet and/or refer to Abcam s IHC antigen retrieval protocol ( Check solution for appropriate color change. (See Technical Notes) 8.6. Gently rinse by gradually adding DI water to the solution, then remove slides and rinse with DI water. Technical Notes: Concentrated Universal HIER antigen retrieval reagent (10X) is a bright yellow color. The RTU or 1X solution is a pale yellow color. When the solution reaches C, the solution turns yellow and indicates that the high temperature solution is at correct ph. Should the ph rise above 7.0, the solution turns a fuschia red color. Should the ph drop too low, the solution turns a pink color. Use positive charged slides and cut tissues into 4-5 micron sections. Do not use any adhesives in the water bath. Poor fixation and processing of tissues will cause tissue sections, especially fatty tissues such as breast, to fall off the slides. Tissues should be fixed for a minimum of 6-12 hours. Protocol time and temperatures for HIER can vary depending on the heat source used. ab Universal HIER antigen retrieval reagent (10X) 6

9 RESOURCES DATA ANALYSIS 9. NOTES ab Universal HIER antigen retrieval reagent (10X) 7

10 RESOURCES ab Universal HIER antigen retrieval reagent (10X) 8

11 RESOURCES ab Universal HIER antigen retrieval reagent (10X) 9

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