How To Use GSIS Method To Detect Cell Insulin Secretion

Min6 Cells Were Selected By Many Customers In This Gift Giving Activity. More Than 50 Cells Were Sent Out, Followed By The Insulin Test. Many Customers Found Only Low Expression& Nbsp; Three Points Should Be Paid Attention To In Min6 Cell Culture: 1. The Cells Are Easy To Agglomerate, And The Digestion Time Seems To Be Very Short, About 30 Seconds, They Will Fall Off Completely. After Falling Off, Please Extend The Digestion Time To 2 Minutes, And Then Neutralize, And The Cell Aggregation Will Be Reduced In The Late Stage. 2. Living Cells Will Float During Transportation. Please Collect The Culture Medium For Centrifugation. After Centrifugation, Lay The Plate Again Until The Complete Culture Medium Has Just Passed Through The Cells. If Too Much Complete Culture Medium Is Added, The Cells Will Become Too Buoyant And Will Not Adhere To The Wall. 3. The Culture Medium Must Be Added With 0.05mm β- Mer  The Following Are The Buffers And Protocols Used In Our Detection. The Steps Must Be Taken Step By Step& Nbsp; Buffer Preparation: NaCl 114mM, KCl 4.7mM, KH2PO4 1.2mM, MgSO4-7H2O 1.16mM, NaHCO3 25.5mM, CaCl2 2.5mM, HEPES 20mM, 0.2% BSA ---> PH 7.2-7.5 (be Sure To Control The PH Range); Operation Steps: Use 2mM Glucose; And 25mM Glucose; As An Example, There Are About 2000 Cells Per Hole In The Cell Planking, And Inactivated Serum Is Used To Remove Antibiotics About Three Days Before Planking+ β- Mer Culture 1640+10% FBS (inactivated)+0.05mm; β- Mer； Incubate In 96 Well Plate For 24 Hours, And Then Change The Medium To 2mM Glucose Medium Overnight. 1. Remove The Medium And Wash Cells With 1 X PBS For 5 Minutes 2. Basic Buffer (2mM  Glucose);, 2 Hours Incubation  --& Gt;& Nbsp; Culture 3. Remove The Supernatant& Gt; 1 X Pbs  Please Be Careful Not To Touch The Bottom During Cleaning 4. Glucose Stimulation (2mM);, Incubation For 1 Hour 5. Harvest Supernatant& Gt; 1 X Pbs  Please Do Not Touch The Bottom During Cleaning. 6. Glucose Stimulation (2mM);, 1 Hour Incubation; 7. Harvest Supernatant& Gt; 1 X Pbs  Please Be Careful Not To Touch The Bottom During Cleaning. 8. Supernatant Sampling& Gt; 1500 Rpm, 5 Min, 4 Degrees; 9. Cell Lysis ---->& Nbsp; Protein Quantification ---->& Nbsp; Hands On  The Following Are The Test Results: Note: To Ensure That The Insulin Secreted By Cells Is Mixed With The Supernatant Sample, The Cells Can Be Gently Stirred On A 100-300 Turn Flat Plate Oscillator During Culture.