You are here: Home > FAQ

 

FREQUENTLY ASKED QUESTIONS
 
 
Areas of interest:
 

 

General Ordering
 
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 ecommerce@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.
 
 
 
 

Our PhaseZERO® team has compiled a list of frequently asked questions by our customers. Many questions posed can be answered within Asterand’s Gene Expression Profiling Methodology document. The FAQs listed below are a quick reference for your questions. If you cannot find an answer to your question here, please contact us so that we may provide you with the information you need.

 

How do you define "normal" tissues?
 
Asterand defines "normal" tissues as those which have been assessed and deemed non-diseased or normal by qualified pathologists. Although a tissue may be classified as normal, it should be noted that the donor may have suffered from a disease that does not explicitly affect the pathological status of the tissue.
  
Why only 3 donors per tissue?
 
The purpose of XpressWay® Profiles is essentially to provide a “snap-shot” of the expression of a gene across the major organs/tissues of the human body. This design allows confirmation or otherwise of a target gene(s) in specific human tissues and, as such, are a first step approach to target validation.

 

 
What are the units of measurement?
 
The unit of measurement is copy number per 100ng total RNA, which has been extrapolated from Asterand’s Global Standard Curve using the qPCR Ct value.

 

What do you use for a housekeeping gene / controls?
  
The target gene is multiplexed with GAPDH in all of the samples to attain confirmation that the reverse transcription and qPCR processes have worked. Also included in each assay are no template controls (NTCs).

 

How do you normalize the data?
 
Asterand’s PhaseZERO™ team does not normalize the data. GAPDH is multiplexed with the target gene in order to confirm that the reverse transcription and qPCR processes have worked. However, many studies have shown that GAPDH is a poor house-keeping gene and, in fact, normalisation of qRT-PCR data using a single house-keeping gene is not considered good practice.

 

How do you reference the data?
 
The data is checked against publically available data. However, no assumptions are made regarding the accuracy of publically available data.

 

What will the reports will look like?
 
An example report is provided on the XpressWay® Profiles web page, or you may click here to view the sample report now.

 

What do the acronyms mean on the report?
 
All terms and acronyms used in the reports are defined in each report on the glossary page at the end of the report

 

The plot shows each value as a single value, do you have the standard deviations since you are testing an n=3?
 
No, because copy number does not exhibit a normal Gaussian distribution and so no statistical analyses are performed using copy number values. Logarithmic transformation of copy numbers allows standard statistical tests to be done.

 

Why do "beach ball" patterns exist?
 
Beach ball patterns indicate that the target is ubiquitously expressed in the tissues included in the XpressWay® Profile.

 

Can you do splice variants/have you done splice variants?
 
Our PhaseZERO® team is highly experienced in providing splice variant data.

 

Can you compare against disease models?
 
There are no data from diseased tissues in our XpressWay® Profiles. However, you can compare the XpressWay® Profile data with your own or published diseased model data or you can request Asterand undertake a CustomMapping project to assess the expression of your target/s of interest in normal and diseased tissues.

 

Can you compare to animal models?
 
There are no data from animals in our profiles. However, you may compare the XpressWay®Profile data with your own or published data.

 

Can you profile on other tissues?
 
Yes, Asterand can provide a CustomMapping project to meet your needs and requirements.

 

How long to do a “new” profile?
 
New profile generations take approximately 4-6 weeks from receipt of an order.

 

How long does it take to get the data on an existing profile?
 
Please allow 2-3 days from receipt of an order to be completely processed and emailed to you.

 

How is the data delivered (electronic, mail, both etc)?
 
Profile data is delivered via e-mail as a PDF version of the report and as an ExcelTM file containing all the data and the donor details.
 
 
 

 
 
 
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 164
0 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 herefor 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 herefor 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?

Yes on a custom isolation. Please contact customerservice@asterand.com for further information.

 
12. How many primary cells are supplied per vial?
 
All Carcinoma Associated Fibroblasts are provided in ampoules of atleast ~1x106 cells.
 
All Chondrocytes and Synovial Fibroblasts are provided in ampoules of ~1x106 cells.
 

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.