Tel:4001790116,13012892256,17521589219

Cell, culture reagent, life science research overall service provider!

Technical Support技术支持

CONTACT US

Isolation And Extraction Of Astrocytes

source:Qida organism  views:625  time:2022-12-16

Astrocytes Are Referred To As Astrocytes And Astrocytes. Astrocytes Are A Kind Of Astrocytes That Exist In The Brain And Spinal Cord In A Star Shape. They Perform Many Functions, Including Biochemical Control Of Endothelial Cells That Form The Blood Brain Barrier, Providing Nutrition To Neural Tissues, Maintaining The Balance Of Extracellular Ions, Regulating Cerebral Blood Flow (CBF), And Repairing Brain And Spinal Cord Injuries Caused By Infection Or Physical Injury It Plays An Important Role In The Formation Of Glial Scar. For How To Separate Astrocytes, Please Refer To The Separation Steps Of Rat Derived Astrocytes Prepared By Shanghai Qida Biology: Prepare Experimental Consumables: Various Gun Heads, Electric Pipettes, Pipettes, 50ML Centrifuge Tubes, 100 Micron Cell Filters, Culture Bottles/dishes, Filter Papers, Ophthalmic Scissors, Surgical Blades, Centrifuges, Fluorescent Microscopes, AGM Astrocyte Culture Medium, GFAP Antibodies, PBS, L-polylysine Penicillin Streptomycin, 0.05% EDTA Trypsin. Begin To Enter The Theme Experiment: Put The Newborn Rats Within 24 Hours On Ice, And Anesthetize Them For 5 Minutes At Low Temperature; Immerse It In 75% Alcohol For 5 Minutes, Cut Off The Head And Put It To Death. Cut The Skin, Subcutaneous Tissue And Skull From Back To Front Along The Middle Of The Brain To The Center Of The Eyebrows. Open The Skull From Both Sides By Ophthalmology, Take Out The Brain Tissue Completely, Place It On The Filter Paper, Cut Off The Cerebellum And Medulla Oblongata With The Surgical Blade, Cut Off The Cerebral Hemispheres On Both Sides Along The Middle Of The Brain, And Remove The Midbrain And Hippocampus. The Blade Moves The Cerebral Cortex To Roll It Around On The Filter Paper To Remove The Cerebral Pia Mater, And Then The Cerebral Cortex Tissue Is Placed In The Cold PBS For Cleaning Three Times. The Liquid Is Sucked Up. ① In 1ml Incomplete Culture Medium, Ophthalmic Scissors Repeatedly Cut The Cortical Tissue Block For About 50 Times To Make It Into Small Pieces Of About 1mm In Size. ② Add 15ml Trypsin Containing 0.05% EDTA, Digest At 37 Degrees For 20 Minutes, And Add Incomplete Medium Twice The Volume To Terminate Digestion. ③ Vortex At 3000r/min For 60s, Centrifugate At 500G For 10min, Discard The Supernatant, Add 30ml Of Medium And Blow Repeatedly To Resuspension The Sediment. ④ Centrifuge 300G For 10 Minutes, Discard The Supernatant, Add 30ml Culture Medium And Blow Repeatedly To Make It Suspend Again. ⑤ Repeat Step 4 Three Times. ⑥ After Filtering Through 100 Mesh Cell Filter Screen, Adjust The Cell Concentration And Plant It In A 75cm2 Culture Bottle Or 25cm2 Culture Bottle Coated With Polylysine (50ug/ml). Change The Liquid In Full Every Two Days, Rinse The PBS Twice Every Time From The Second Time, And Shake The Culture Bottle Horizontally For About 50 Times Each Time& Nbsp; When The Cell Growth Density Is Above 80%, Immunofluorescence Identification Is Carried Out. Generally, GFAP-FITC Immunofluorescence Combined With DAPI Staining Is Used To Identify The Purity Of Astrocytes& Nbsp; That Is, Astrocyte Purity=GFAP Positive Cells/DAPI Positive Cells& Nbsp; Small Details Determine The Success Or Failure Of The Experiment. The Following Points Need More Attention: 1. No Matter What Tissues Are Separated, Pay Attention To Selecting Fresh Tissues (no More Than 24 Hours), And Try To Select Rats And Mice Within Three Days Of Birth. The Sampling Process Should Be As Sterile As Possible, And The Meninges, Blood Vessels And Hippocampus Should Be Removed As Much As Possible. 2. Attention Shall Be Paid To The Elimination Of Residual Blood On The Surface Of Brain Tissue As Far As Possible To Avoid The Influence Of Blood Cells And Some Serum Components On The Adhesion Of Glial Cells. 3. The Digestion Concentration And Time Of Enzyme Should Be Strictly Controlled. 4. Strictly Control The Conditions Of Glial Cell Culture, Including The Quality And Concentration Of Cell Culture Medium (culture Medium, Growth Factor, Serum, Antibiotic). Qida Biological Astrocyte Culture Medium AGM (Qida Biological, Product No.: P2701) Can Be Directly Selected To Save Time And Worry. The Finished Product Can Be Prepared And Directly Added. 5. Polylysine Is Selected As The Coating Solution With A Concentration Of 25-50ug/ml. Because Of The Chemical Nature Of Polylysine, Excessive Use Will Lead To Cell Death
×