Method for inducing and acclimating epidermal stem cells into nerve cells
A technology of epidermal stem cells and nerve cells, which is applied in the field of using epidermal stem cells to induce domestication into nerve cells, and can solve the problems of limited application scope of epidermal stem cells.
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0035] Example 1 Preparation of epidermal stem cells before induction and domestication
[0036] 1. Preparation of epidermal stem cells:
[0037] Direct source and original source: With the consent of the patient, it was taken from a healthy circumcision patient in the 301 Hospital Surgical Clinic.
[0038] (1) Take fresh adult foreskin (isolated for about 1 to 4 hours, stored at 4°C);
[0039] (2) Soak the foreskin in a dilute iodophor solution (containing 20% iodophor) for 1 min;
[0040] (3) PBS / physiological saline rinse 3 times to wash away blood clots (to prevent affecting enzyme digestion);
[0041] (4) Trim the foreskin, remove the fat tissue, and then rinse with PBS / saline several times, and finally cut into 1cm 2 The left and right organizational blocks;
[0042] (5) Put the tissue block in a 6cm dish containing a solution of neutral protease (3mg / ml; D-hanks as solvent) and treat it at 4°C for 14-16h. If the treatment is not enough, place it in a 37°C incubator (or directly ...
Embodiment 2
[0118] Example 2 Directed induction and domestication of epidermal stem cells into nerve cells
[0119] 1. Medium composition
[0120] (1) One component and dosage of nerve induction medium are:
[0121] DMEM / F12 (Invitrogen, cat.no.11330-32); 2% FBS (Gibco, cat.no.10437010) 45ng / ml b-FGF (Invitrogen, cat.no.13256-029); 20ng / ml EGF( invitrogen, cat.no.); 0.5mM retinoic acid(RA)(sigmaR2625); 0.5mM valproic acid(Sigma, cat.no.T8552); 2000U / ml leukemiainhibitory factor(Chemicon, cat.lif1010); 100mM sodium pyruvate( sigma, cat.no.P2256); 0.5mM β-mercaptoethanol(sigma cat.no.m7522);
[0122] (2) The two components and dosage of nerve induction medium are:
[0123] DMEM / F12 (Invitrogen, cat.no.11330-32) 2% FBS (Gibco, cat.no.10437010); 20ng / ml b-FGF (Invitrogen, cat.no.13256-029); 20ng / ml EGF( invitrogen, cat. no.); 0.5 uM RA (sigma R2625); 1×B27 (invitrogen Cat. No. 17504); 100 ug / ml butylated hydroxyanisole (sigma, cat. no. B1253).
[0124] (3) The components and dosage of dry maintenance...
Embodiment 3
[0129] Example 3 Identification and result analysis of target cells
[0130] 1. Immunofluorescence identification of induced differentiation results
[0131] 1. Identification method: In the experimental verification of induced nerve cells, the immunofluorescence detection of neural cell marker (a-SMA) identification method is as follows:
[0132] (1) Take three weeks after induction and epidermal stem cells as controls, and wash the cells twice with PBS.
[0133] (2) Add 1ml of Tribule lysis solution pre-cooled at 4℃ to each six-well plate, and see that the cells are all lysed, about 5 minutes
[0134] (3) Use a ribozyme-free gun tip to aspirate the cell lysate into a 1.5ml centrifuge tube
[0135] (4) Add 200ul of chloroform, shake vigorously up and down four to five times to mix well
[0136] (5) 12000rpm, 5min 4℃. The liquid is divided into three layers, suck the upper transparent liquid layer into a new 1.5ml centrifuge tube
[0137] (6) Add an equal volume of isopropanol, mix upside ...
PUM
Abstract
Description
Claims
Application Information
- R&D Engineer
- R&D Manager
- IP Professional
- Industry Leading Data Capabilities
- Powerful AI technology
- Patent DNA Extraction
Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.
© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com