1. What is The Asterand Material Transfer Agreement (MTA)/Terms and Conditions of Sale (T&C)?
Asterand’s Terms and Conditions of Sale govern the relationship between Asterand and its Clients. The Terms and Conditions of Sale address bioethics requirements, methods of ordering, and procedures for returning unsatisfactory samples or data. The Terms and Conditions of Sale also set forth the appropriate method of payment. Terms and Conditions of Sale must be signed by an authorized representative of the entity named as the Client. Where the Client is a university or academic center, please arrange for signature by an individual having authority to sign on behalf of the university or academic center. Asterand may accept orders for all researchers at that university or academic center under one Terms and Conditions of Sale, provided it has been approved at the entity, and not department, level.
2. How can I order items that are not available on-line?
Please send an email to customerservice@asterand.com outlining what you require for your research, all relevant contact
of the institution/company where you conduct your research, and details about the samples that you require
(e.g. tumor with adjacent normal, female donors only, age ranges, and the like). We will follow up with you and connect you with a Regional Director of Business Development for your area.
3. How can I make payment for my order?
Please refer to our How to Order page.
Primary Cells and Cell Lines
1. What type of growth media do you recommend for cells and cell lines?
a. Chondrocytes and Synovial Fibroblasts are grown in DMEM: F12 with 10% FBS
b. Stromal Fibroblasts are grown in DMEM with 10% FBS
c. Epithelial cells are grown in keratinocyte serum free medium from gibco containing 2% FBS and supplement provided by Gibco.
d. The WSU cell lines are grown in RPMI 1640 with Phenol Red with 10% FBS
e. The MCF10DCIS.com cell line is grown in DMEM:F12 with 5% horse serum.
f. The SUM cell lines are grown in Ham’s F12 and each have their own recipe of supplements for optimal growth.
click here for more details.
g. The Onyvax (OPC) lines are grown in keratinocyte serum free medium from gibco containing 2% FBS and supplement provided by Gibco.
click here for more details.
2. How should I freeze cell lines?
Whenever you use your own media for freezing we recommend freezing only a couple of vials, then thaw one of them to make sure your cells will grow again. We recommend the cryostor because you will get higher viability upon thawing. We use a freezing medium by BioLife Solutions called crystor CS5. We have very high viability upon thawing when using this media. biolifesolutions.com/products/cryopreservation.htm
3. Is it possible to use the cells to make xenograft on nude mouse by injection?
The following cell lines have been used in mice: SUM149, SUM159, SUM225, SUM1315, MCF10DCIS.com
4. What storage temperatures do recommend for cell lines and primary cells?
Short term storage can be at -80 degrees celcius. If longer than 2 days, all cells should be stored in vapor phase liquid Nitrogen storage.
5. How is the tissue for isolation of primary cells obtained?
All cells are obtained from post-mortem tissue with the exception of knee synovial fibroblast and chondrocytes. They are from either surgical or post mortem. This information is included in the associated clinical data.
6. Are your Human Chondrocytes and Fibroblasts available in proliferating form (ie. in media) or are they all in frozen ampoules?
We can provide them in proliferating format but there will be an additional cost.
7. How are primary cells characterized/phenotyped? Do you perform immunocytochemistry for smooth muscle actin or FAP (fibroblast activation protein) or any other marker?
Stromal fibroblasts were characterized by IHC using vimentin and visually confirmed as fibroblasts.
Synovial fibroblasts were characterized by IHC using vimentin and collagenase I. Chondrocytes were characterized by IHC using aggrecan and collagenase II. Epithelails were characterized by IHC using Cyotkeratin 18.
8. If so, what percentage of cells is actually positive for these markers, on average?
Upon visual inspection of the IHC stained cells, the cells generally stain 95-99% positive for the different markers.
9. How many passages do you recommend for primary cells?
We recommend using the primary cells between passages 5 and 10.
10. Do you have a protocol for immortalization?
All immortalized cell lines were spontaneously immortalized by the researchers who provided them. We do not have a protocol for immortalizing cells.
11. Is RNA available for these cell lines?
Serum
1. When was the blood drawn?
The blood draw is pre-surgery and post-cancer diagnosis.
2. Is the serum from a single donor? Or, has it been pooled?
The serum available on-line is from a single donor.
3. Is plasma available?
In many cases plasma is available from the donor that provided the serum. Please send an email to customerservice@asterand.com with the serum sample IDs that you would like us to match with available plasma.
RNA
What method do you use to quantify RNA?
We quantify RNA and DNA with the NanoDrop spectrophotometer. Our current protocol for extracting RNA is a modified organic extraction plus an on column DNase digestion step. The RNA is eluted in 0.1mM EDTA in DEPC water and stored in a -80° freezer (short-term) & -180° freezer (long term).
