大鼠脑周细胞Cell Specification
Brain Vascular Pericytes (BVP) are perivascular cells that are closely associated with the
endothelium of capillaries and other small vessels [1] BVP, located between endothelial cells
and astrocytes in the brain communicate with other cells by extending long cytoplasmic
processes which wrap around the capillaries [1, 2 BVP participate in a variety of processes
including angiogenesis, endothelial cell survival, regulation of capillary blood flow, and
establishment and maintenance of the blood brain barrier [3, 4 Pericyte dysregulation has been
linked to several pathological conditions such as hypertension, diabetic retinopathy,
atherosclerosis, multiple sclerosis, Alzheimer’s disease, and tumor angiogenesis [2, 4]. The
unique and diverse functions of BVP make them novel candidates for cell therapy in regenerative
medicine. Cultured rimary rat BVP (RBVP) are a useful in vitro model for understanding the
molecular mechanisms of blood brain barrier regulation and for studying a wide variety of
central nervous system diseases
BVP from ScienCell Research Laboratories a e isolated from adult rat brain BVP are
cryopreserved at passage one and delivered frozen. Each vial contains >5 x 10 cells in 1 ml
大鼠脑周细胞volume. BVP are characterized by immunofluorescence with antibody specific to platelet
derived growth factor  (PDGF receptor  and  smooth muscle actin. BVP are negative for
mycoplasma, bacteria, yeast, and fungi. BVP are guaranteed to further expand for 5 population
doublings under the conditions provided by ScienCell Research Laboratories.
Recommended Medium
It is recommended to use Pericyte Medium PM, Cat. #1201 for culturing BVP in vitro.
Product Use
BVP are for research use only. They are not approved for human or animal use, or for
application in in vitro diagnostic procedures.
Storage
Upon receiving, directly and immediately transfer the cells from dry ice to liquid nitrogen and
keep the cells in liquid nitrogen until they are needed for experiments
Shipping
Dry ice.
References
[1] Dore Duffy P, Cleary K. (2011) “Morphology and properties of pericytes.” Methods Mol Biol. 686:49 68.
[2] Allt G Lawrenson JG. (2001) “Pericytes: cell biology and pathology.” Cells Tissues Organs 169: 11.
[3] Daneman R, Zhou L, Kebede A, Barres B. (2010) “Pericytes are required for blood brain barrier integrity during
embryogenesis.” Natur . 468:562 566.
[4] Kutcher M, Herman I. (2009) “The pericyte: cellular regulator of microvascular blood flow.” Microvasc Res. 77:
235 246.
Instructions for culturing cells
Caution: Cryopreserved cells are very delicate. Thaw the vial in a 37 C water bath
and return the cells to culture as quickly as possible with minimal handling!
大鼠脑周细胞Initiating the culture:
1. Prepare a poly lysine coated culture vessel (2 μg/cm , T 75 flask is recommended).
Add 10 ml of sterile water to a T 75 flask and then add 15 μl of poly lysine stock
solution (10 mg/ml, Cat. #0413). Leave the vessel in incubator overnight (minimum one
hour at 37 C incubator).
2. Prepare complete medium. Decontaminate the external surfaces of medium bottle and
medium supplement tubes with 70% ethanol and transfer them to a sterile field.
Aseptically transfer supplement to the basal medium with a pipette. Rinse the tube with
medium to recover the entire volume.
3. Rinse the poly lysine coated vessel with sterile water twice and then add 15 ml of
complete medium. Leave the vessel in the sterile field and proceed to thaw the
cryopreserved cells.
4. Place the frozen vial in a 37 C water bath. Hold and rotate the vial gently until the
contents completely thaw. Remove the vial from the water bath promptly, wipe it down
with 70% ethanol and transfer it to the sterile field.
5. Remove the cap carefully without touching the interior threads. Gently resuspend and
dispense the contents of the vial into the equilibrated, poly lysine coated culture vessel.
A seeding density of 5,000 cells/cm is recommended.
Note: Dilution and centrifugation of cells after thawing are not recommended since these
actions are more harmful to the cells than the effect of residual DMSO in the culture. It is
also important that cells are plated in poly lysine coated culture vessels to promote cell
attachment.
6. Replace the cap or lid, and gently rock the vessel to distribute the cells evenly. Loosen
cap if necessary to allow gas exchange.
7. Return the culture vessel to the incubator.
8. For the best result, do not disturb the culture for at least 16 hours after the culture has
been initiated. Refresh culture medium the next day to remove the residual DMSO and
unattached cells
Maintaining the culture:
1. Refresh supplemented culture medium the next morning after establishing a culture from
cryopreserved cells.
2. Change the medium every three days thereafter, until the culture is approximately 70%
confluent.
3. Once the culture reaches 70% confluency, change medium every other day until the
culture is approximately 90% confluent.
Subculturing:
1. Subculture when the culture reaches 90 95% confluency
2. Prepare poly lysine coated culture vessels (2 μg/cm one day before subculture.
3. Warm complete medium, trypsin/EDTA solution (T/E, Cat. #0103), T/E neutralization
solution (TNS, Cat. #0113), and DPBS (Ca
++
and Mg
++
free, Cat. #0303) to room
temperature. We do not recommend warming reagents and medium at 37 C water bath
prior use.
4. Rinse the cells with DPBS.
5. Add 8 ml of DPBS and then 2 ml of T/E solution into flask (in the case of T 75 flask).
Gently rock the flask to ensure complete coverage of cells by T/E solution. Incubate the
flask at 37 C incubator for 1 to 2 minutes or until cells completely round up. Use
microscope to monitor the change in cell morphology.
6. During incubation, prepare a 50 ml conical centrifuge tube with 5 ml of fetal bovine
serum (FBS, Cat. #0500).
大鼠脑周细胞7. Transfer T/E solution from the flask to the 50 ml centrifuge tube (a few percent of cells
may detached) and continue to incubate the flask at 37 C for another 1 to 2 minutes (no
solution in the flask at this moment).
8. At the end of incubation, gently tap the side of the flask to dislodge cells from the
surface. Check under microscope to make sure that all cells detach.
9. Add 5 ml of TNS solution to the flask and transfer detached cells to the 50 ml centrifuge
tube. Rinse the flask with another 5 ml of TNS to collect the residual cells.
10. Examine under microscope for a successful cell harvest by looking at the number of cells
being left behind. There should be less than 5%.
Note: Use ScienCell T/E solution that is optimized to minimize cell damages due to over
trypsinization.
11. Centrifuge the 50 ml centrifuge tube at 1000 rpm for 5 min. Resuspend cells in culture
medium.
12. Count and plate cells in a new, poly lysine coated culture vessel with cell density
as recommended.
Caution: Handling animal derived products is potentially biohazardous. Always wear gloves
and safety glasses when working with these materials. Never mouth pipette. We recommend
following the universal procedures for handling products of human origin as the minimum
precaution against contamination [1].
[1] Grizzle , Polt S. (1988) “Guidelines to avoid personal contamination by infective agents in research
laboratories that use human tissues.” J Tissue Cult Methods. 11: 191 9.