Collection - Cellular Immunofluorescence Climbing Procedure

  In Mammalian Cell Research, It Is Often Necessary To Do Immunofluorescence Staining Of Cells. Qida Biology Has Sorted Out The Operation Steps Of Cellular Immunofluorescence Slides For You, And Praised The Collection. The Next Experiment Will Not Be In A Hurry. DAY 1:1. Clean The Cover Glass With Detergent, Then Rinse The Detergent With Tap Water, Rinse It With Pure Water For Three Times, And Soak It In 75% Alcohol For 10min. 2. Use Tweezers To Pick Up A 24mm * 24mm Cover Glass And Burn The Alcohol Dry On The Alcohol Lamp. The Temperature Should Not Be Too High. 3. Put The Dried Cover Glass Into A Six Hole Plate, And Then Cool The Cell Suspension (about 1-2 × 104 Cells) On The Cover Slip. After 4.5h, Slightly Add 1ml Of Medium, And Put It Into A 37 ℃ 5% CO2 Incubator For Overnight Culture. The Cells Grow Well On The Slide& Nbsp;& Nbsp; Humsc Cell Staining (Qida Biological, Product No.: CD5007) DAY 2:1 In The Culture Plate, The Slides With Cells Have Been Soaked With PBS For 3 Times, 3 Minutes Each Time& Nbsp; 2. Fix The Slide With 4% Paraformaldehyde For 15min, And Soak The Slide With PBS For 3 Times, 3min Each Time& Nbsp; 3. 0.5% Triton X-100 (prepared With PBS) Is Permeable For 20 Min At Room Temperature (this Step Is Omitted For Antigen Expressed On Cell Membrane)& Nbsp; 4. PBS Was Immersed In The Glass Slide For 3 Times, 3 Min Each Time. PBS Was Dried With Absorbent Paper, And Normal Goat Serum Was Dropped On The Glass Slide, And The Glass Slide Was Sealed At Room Temperature For 30 Min& Nbsp; 5. Absorb The Sealing Solution With Absorbent Paper And Do Not Wash It. Add Enough Diluted Primary Antibody To Each Slide And Put It Into A Wet Box. Incubate At 4 ℃ Overnight; DAY 3：6. Add Fluorescent Secondary Antibody: PBST Was Soaked And Washed For 3 Times, 3 Minutes Each Time. After Absorbing The Excess Liquid On The Climbing Piece With Absorbent Paper, The Diluted Fluorescent Secondary Antibody Was Dripped And Incubated In A Wet Box At 20-37 ℃ For 1 Hour. PBST Was Soaked And Washed For 3 Times, 3 Minutes Each Time; Note: From The Addition Of Fluorescent Secondary Antibody, All The Following Operation Steps Should Be Carried Out In A Dark Place As Far As Possible& Nbsp; 7. Re Staining Nucleus: Add DAPI Dropwise And Incubate It In Dark For 5min, Dye The Nucleus On The Specimen, And Wash Off The Excess DAPI With PBST 5minx4 Times; 8. Absorb The Liquid On The Climbing Piece With Absorbent Paper, Use The Sealing Liquid Containing Anti Fluorescence Quenching Agent To Seal The Piece, And Then Observe And Collect The Image Under The Fluorescence Microscope. Precautions: 1. When Taking The Cell Slide, The Action Should Be Gentle To Prevent Crushing The Cell Slide And Affecting The Experimental Process& Nbsp; 2. In The Process Of Seeding Cells, The Cells Should Be Gently Mixed And Shaken In The Shape Of "eight" Or "cross" To Prevent The Cells From Growing Too Densely Locally.