大鼠双丝氨酸/苏氨酸酪氨酸蛋白激酶(DSTYK) ELISA Kit

MATERIALS REQUIRED BUT NOT SUPPLIED

• Microplate reader with 450 ± 10nm filter.
• Precision single or multi-channel pipettes and disposable tips.
• Eppendorf Tubes for diluting samples.
• Deionized or distilled water.
• Absorbent paper for blotting the microtiter plate.
• Container for Wash Solution.

 
 STORAGE OF THE KITS

• For unopened kit: All the reagents should be kept according to the labels on vials. The TMB Substrate, Wash Buffer (30 × concentrate) and the Stop Solution should be stored at 4^oC upon receipt while the others should be at -20^oC.
• For opened kit: When the kit is opened, the remaining reagents still need to be stored according to the above storage condition. Besides, please return the unused wells to the foil pouch containing the desiccant pack, and reseal along entire edge of zip-seal.

 
Note:
It is highly recommended to use the remaining reagents within 1 month provided this is within the expiration date of the kit. For the expiration date of the kit, please refer to the label on the kit box. All components are stable until this expiration date.
 
 SAMPLE COLLECTION AND STORAGE

• Serum - Use a serum separator tube and allow samples to clot for two hours at room temperature or overnight at 4^oC before centrifugation for 20 minutes at approximately 1000×g. Assay freshly prepared serum immediately or store samples in aliquot at -20^oC or -80^oC for later use. Avoid repeated freeze/thaw cycles.
• Plasma - Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples for 15 minutes at 1000×g at 2 - 8^oC within 30 minutes of collection. Remove plasma and assay immediately or store samples in aliquot at -20^oC or -80^oC for later use. Avoid repeated freeze/thaw cycles.
• Tissue homogenates - The preparation of tissue homogenates will vary depending upon tissue type. For this assay, tissues were rinsed in ice-cold PBS(0.01mol/L,pH 7.0-7.2) to remove excess blood thoroughly and weighed before homogenization. Minced the tissues to small pieces and homogenized them in 5-10 mL of PBS with a glass homogenizer on ice(Micro Tissue Grinders woks, too). The resulting suspension was sonicated with an ultrasonic cell disrupter or subjected to two freeze-thaw cycles to further break the cell membranes. After that, the homogenates were centrifugated for 5 minutes at 5000×g. Remove the supernate and assay immediately or aliquot and store at ≤-20^oC.  
• Cell Lysates - Cells must be lysed before assaying according to the following directions.   

• Adherent cells should be detached with trypsin and then collected by centrifugation (suspension cells can be collected by centrifugation directly).
• Wash cells three times in cold PBS.
• Resuspend cells in PBS (1×) and the cells was subject to ultrasonication for 4 times (or Freeze cells at ≤-20^oC. Thaw cells with gentle mixing. Repeat the freeze/thaw cycle for 3 times.). 
• Centrifuge at 1500×g for 10 minutes at 2 - 8^oC to remove cellular debris.

• Cell culture supernates and other biological fluids - Centrifuge samples for 20 minutes at 1000×g. Remove particulates and assay immediately or store samples in aliquot at -20^oC or -80^oC. Avoid repeated freeze/thaw cycles.

 
Note:

• Samples to be used within 5 days may be stored at 4^oC, otherwise samples must be stored at -20^oC (≤1 month) or -80^oC (≤2 months) to avoid loss of bioactivity and contamination.
• Sample hemolysis will influence the result, so hemolytic specimen can not be detected.
• When performing the assay, bring samples to room temperature.