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Growth Factors

Posted by Karen on September 9th, 2015  ⟩  0 comments

Bovine Serum Albumin (BSA) comes in many forms, which is why people frequently post questions online, asking what BSA should be used in their experiment. Forum responses have been good about directly addressing the procedure in question, but it's really hard to find a consolidated resource that helps researchers evaluate each type of BSA and their appropriate uses. Since the Internet is lacking a central guide, I set out to investigate what goes into choosing between BSA types and provide these answers in this article.

First, we should address what types of BSA exist in the market. GoldBio offers two types: protease free and fatty acid free (I’ll explain why fatty acid free is a good choice for many experiments later on in this article). But then you have other types of BSA, such as standard BSA, low endotoxin, immunoglobulin free and blocking agent BSA. Obviously each type offers certain features that may help or hurt your project. So which one is best for your experiment?

To figure this out, we should look at common processes that use BSA:

1.Cell Culture

2.ELISA

3.Enzyme System Diluent

4.Immunology

5.Protein Stabilization

6.Carrier Protein



One Chart to Rule Them All

Which one of these six common methods are you using? That’s going to be what helps you decide which BSA is right for your experiment. Certain types of BSA will cover a multitude of techniques; for that reason, it is better to address which BSA types are appropriate for each process in a simple chart rather than long paragraphs where overlaps aren't as evident.
BSA Selection Guide Chart - What BSA (Bovine Serum Albumin) Should I Use – Your BSA Selection Guide






  

Beyond the Chart:

The chart certainly reduces having to do heavy research into finding the best option. It also makes it clear that there are certain BSA types that reach further than others. For example, it's obvious that fatty acid free BSA and protease free BSA works well in a wide variety of techniques, making each a more ideal product, while a blocking BSA is best used only as a blocking agent in ELISA. 

But beyond these criteria, there are other very important factors to consider. Number one is usually price. Being sure it fits the budget is make-or-break for almost every researcher. The next go-to for a researcher is purity. But let’s say you found the right BSA type, it fits your budget and it has excellent purity, what else should you consider? When evaluating BSA, the location it’s manufactured in and if it’s certified as bovine spongiform encephalopathy compliant is also vital for obvious reasons.







Why Choose Fatty Acid Free BSA?

The factors mentioned in this article are exactly what GoldBio considered when first offering protease free BSA, and then more recently our fatty acid free BSA, which is also protease free. We were determined to provide researchers with a product that satisfies most criteria for BSA selection. First, they’re both extremely affordable and have high purity. Beyond that, however, both products are manufactured in the USA and are BSE/TSE compliant. And while GoldBio’s protease free BSA is appropriate for a variety of common techniques, we recently introduced fatty acid free BSA because it is useful in an even broader range of experiments, not just the ones listed on the chart. Besides satisfying some check boxes, fatty acid free BSA eliminates certain variables from the experiment. In fact, there are times when the nature of the experiment necessitates the use of fatty acid free BSA over others. For instance, if you're working with a fatty acid sensitive cell culture system such as CHO, Vero and MDCK cells, fatty acid free BSA is more suitable for those situations. 

Of course both products have their limits. They won't be ideal for processes such as PCR or bloodbanking, but in general, both BSA types, and more particularly the fatty acid free BSA will allow you to stretch the product over a wider variety of techniques without costing much more. Economically, it's a better choice. And if you still question whether this product or other BSA types are right for you, it’s best to do additional research to be certain. 




If you found this guide useful, you might want to share it with others and check out some of our other articles, which are both helpful and entertaining. 







    
              Karen Martin
GoldBio Marketing Coordinator


"To understand the universe is to understand math." My 8th grade
math teacher's quote meant nothing to me at the time. Then came
college, and the revelation that the adults in my past were right all
along. But since math feels less tangible, I fell for biology and have
found pure happiness behind my desk at GoldBio, learning, writing
and loving everything science. 



Category Code: 88253 79105

Posted by Karen on August 14th, 2015  ⟩  1 comments

It’s time to choose a cell line for your project. Research has led you to a few candidates, but narrowing them down can be difficult and the decision goes beyond what works for your experiment. Genetics, identification, sources, reproducibility and a whole lot more are critical characteristics to consider. To help simplify this not so simple process, I’ll break it down into a few topics that you should consider when choosing or working with a particular cell line.

How to Really Choose a Cell Line - From the Basics to the Complex Aspects of Deciding on Your Cell Line

The Basics for Choosing Your Cell Line:

The immediate criteria you consider are: How easy is the cell line to work with? How fast will it grow? And how accessible is it? But next on the list of things to consider, which is critical, is making sure the cell line you choose best mimics the traits you’re trying to study. Does it grow fast or slow? Does it spread easy or not? What do the publications say about each cell line of interest, and which one best represents the work you’re trying to do?


Going Slightly Beyond the Basics:

Your motives are also going to play a role in the cell line you choose. For example, if you’re doing stem cell research, you will need to also consider which cells have a higher differentiation potential for the ectoderm, endoderm or mesoderm. And that will further depend on which cells you want to study. If you want to study kidney cells, you would need to select cell lines that have a higher differentiation potential for the mesoderm.



  


The Complex Aspects of Choosing Your Cell Line:

When doing very specific research, deciding on your cell line is going to require far more variables. If you’re testing the behavior of a drug, selecting a cell line cannot be as simple as saying “Well I’m studying prostate cancer, so I’ll stick with the classic PC-3 line.” According to Aaron Krol’s article “The New Cancer Models, Part 3: Cell Line Critics,” a cell line that has been repeatedly published is often chosen without other more important considerations such as how closely it falls in line with the genetics of the cancer in question. If approaching cell line selection based on what has been popularly published, without doing rigorous research, you might not get the most accurate picture. Instead, you should take more care in examining your choice behind a particular cell line.

Memorial Sloan-Kettering Cancer Center has developed the cBioPortal, which can help make this task a little easier by providing researchers with a way to visualize and analyze cancer genomics data sets. Another program creating user-friendly benefits is Broad Institute’s and Novartis’s Cancer Cell Line Encyclopedia. The CCLE provides genetic characterization of human cancer models. Krol’s article explained that researchers can use the CCLE to measure cell line information against tumor genetic information and essentially score how closely related the cancer is to the potential cell line.



The Misidentification of Cell Lines:

Combing through endless genetic information may seem very tedious, and the scrutiny can’t stop there. When choosing a cell line and planning your purchase, you should evaluate the history of the cell line you’re interested in. Reproducibility (something GoldBio believes in and has written about) is becoming a trending topic within the scientific community, which makes this step necessary; however, it is also very important to be sure of what you’re using because of the impact your research may have on the public. When you’ve identified the cell line you want to work with, it is not always good practice to obtain it from a neighboring lab, the reason being that you have to spend extra time being certain of what you were given. All too often, cell lines are mislabeled. Moreover, you also need to confirm that there has been no contamination, which can happen in any lab. When it comes to very sensitive research, be sure of the company, quality and source of your cell line. Too many cell lines have been misidentified so it’s important to know that a specific, cited cell line has conformed to its same function over the ages.

The International Cell Line Authentication Committee has a database listing cell lines that are known to be cross-contaminated. While this is a helpful tool for identifying cell lines in question, practicing some quality controls within your own lab while working with your cell line is also necessary. 



Beyond Choosing a Cell Line:

There are other questions you might have considered when choosing your cell line. For example, if you’re working with a human cell line cell, will your murine growth factors be compatible? Thankfully, if you shop GoldBio’s wide selection of growth factors, that information will be provided for you, either directly in the product description or in the associated documents, and if you can’t find it, you can always ask. Let’s look at FGF1, Murine. Within the body of the description there is mention that “FGF1 exhibits considerable cross-reactivity…” Furthermore, there is a table below the main description that shows the cross-species homology. You can be assured that flexibility does exist in this example, but this shows you how to double check.




How to Really Choose a Cell Line - From the Basics to the Complex Aspects of Deciding on Your Cell Line




What about storing and reviving your cell line? Animal cell lines can be preserved in either cryogenic freezers or in liquid nitrogen. Due to the expense of cell lines, it is important to be careful when preserving your supply. During the freezing process, temperatures must be lowered slowly. However, when reviving your cells, the thawing process must be rapid. And if you’re working with a more unexplored cell line, you should record the percentage of viable cells using Trypan Blue stain.

When all is said and done, you also want to ensure reproducibility. Even when you have researched the history of your cell line and its genetics, and you purchased your cells from a trusted repository there are more steps to building on and validating your work. While a cell line gives you a great picture of in vitro situations, it’s important to then research primary cells to use in order to test your experiment. Primary cells are not ideal for initial research because they hold some significant limitations such as being slow to grow, having limited life spans and little to compare with. However, they offer the benefit of giving you a better in vivo picture. Therefore, it’s good practice to replicate the results of your experiment in primary cells.



Takeaway:

The takeaway from this article is to be very critical when selecting the right cells. Ask the right question, spend time doing the research, purchase all products from trusted providers (don’t borrow anything for sensitive experiments), be able to replicate and be sure your lab follows the same procedures. Even though this process isn’t standardize, there are some guides available, and there is nothing wrong with developing a standard for your lab based on published resources.



Broad Institute, Novartis Institutes for Biomedical Research, Genomics Institute of the Novartis Research Foundation (2013).     Broad-Novartis Cancer Cell Line Encyclopedia (CCLE). Retrieved August 10, 2015


Gao et al. Sci. Signal. (2013) & Cerami et al. Cancer Discov. (2012). cBioPortal. MSKCC. Retrieved August 10, 2015.

International Cell Line Authentication Committee (October 10, 2014). Database of Cross-contaminated or Misidentified Cell Lines.     Version 7.2. Retrieved August 10, 2015.

Krol, A. (2013). The New Cancer Models, Part 3: The Cell Line Critics. Bio-IT World. Retrieved August 10, 2015.

Yu, M., Selvaraj, S., Liang-Chu, M., Aghajani, S., Busse, M., Yuan, J., . . . Neve, R. (2015). A resource for cell line authentication,    annotation and quality control. Nature, 520(7547), 307-311. doi:10.1038/nature14397




    
              Karen Martin
GoldBio Marketing Coordinator


"To understand the universe is to understand math." My 8th grade
math teacher's quote meant nothing to me at the time. Then came
college, and the revelation that the adults in my past were right all
along. But since math feels less tangible, I fell for biology and have
found pure happiness behind my desk at GoldBio, learning, writing
and loving everything science. 



Category Code: 88253 79107

Posted by Karen on June 16th, 2015  ⟩  0 comments

Given our technology, specifically our reliance on alarms and digital calendars to map out our daily activities, it’s easy to overlook the environmental signals that influence our body’s reaction to the rising and setting of the sun. In fact, it’s because of our technological dependence, and perhaps due to the “Hot Tub” episode from “Seinfeld” (since Kramer can’t be taken seriously), that before my junior or senior year in college, I really never thought that a “mental clock” or the circadian rhythm applied to humans.


Yet, as much as we ignore the role these signals play, they influence hormone release, RNA transcription, body temperature and other functions. Disruptions can cause a lot of immediate reactions such as jet lag, but it is also believed that continued disruption of an individual’s mental clock increases his or her risk for certain cancers.

Researchers have put more effort into understanding the influence and mechanisms behind the circadian clock. For instance, it has been long believed that light quantity, or irradiation, was the primary input for the body’s master clock - the superchiasmatic nucleus (SCN). As light gets brighter, excitation occurs and cells begin to shift into new patterns that prepare for the coming day. Similarly, as the day grows darker, cell patterns and transcription patterns change, and the body begins to prepare for the coming night.

In mammals, the retino-hypothalamic projection, which is composed of a range of retinal ganglion cells, is what delivers visual input to the SCN. These retinal ganglion cells are photosensitive and can have a role in training the circadian clock when rod and cone photoreceptors are not present.

However, since all photoreceptor types are present in this pathway, University of Manchester researcher Tim Brown and his colleagues used this information to form the hypothesis that light quality (color) is another powerful and reliable influencer of the circadian rhythm.

Brown and his team first began research by determining whether changes in colors associated with the earth’s rotation could provide information about the sun’s angle (when viewed from earth) that would be useful for the internal determination of the time of day. Their results showed significant predictability in changes of brightness and color based on the solar angle. 

Your Mental Clock in HD – Circadian Rhythm Improved Color & Brightness - Research Graphs

The next step was proving these changes could be noticed by mammalian visual systems. Using mice, they determined that blue to yellow transitions were more noticeable during twilight hours, meaning color was a factor in discerning night from day.

While it was established that color changes could be detected during twilight hours, and it provided significant information about the time of day, Brown and his team needed to find out if color had a physiological impact on mammals. To do this, they set up an experiment where an artificial sky was displayed to freely moving mice in their cages. When researchers tested irradiance alone, they observed electrophysiological changes; however, when their experiment included color, they found the circadian pattern was more accurately altered. For example, in every case where irradiance was tested alone, the body temperature in mice peaked later than when twilight colors were included.

Brown’s results not only suggest that blue and yellow colors during twilight hours influence the circadian rhythm, but the colors also provide more accurate inputs for the SCN that can quickly and effectively train the body around a diurnal cycle.

The immediate question one might have is whether this applies to nocturnal animals and those with colorblindness. According to Brown, this discovery applies to approximately 90% of mammalian species, which includes both diurnal and nocturnal mammals because they have the capacity for blue-yellow color vision. Therefore, this mechanism is not only found in nocturnal mammals, but it can also work for people with red-green colorblindness (deuteranopia).

Brown also found in his research that clock cells in mice can respond to very momentary color changes, which means that geographic latitudes will not have a significantly negative impact on a mammal’s ability to distinguish time of day.

There is no doubt that the circadian rhythm is loaded with sensitive and perplexing layers for researchers to uncover. While this study highlights another reliable influencer, other recent studies have identified how connected the circadian clock and metabolism are, and furthermore how gut flora also has an influential role. So, for a show about nothing, with quips about exaggerated human events and "yadda yadda yadda," perhaps there is more truth to its humor than we originally thought.



Walmsley, L., Hanna, L., Mouland, J., Martial, F., West, A., Smedley, A., . . . Brown, T. (2015). Colour As a Signal for Entraining the
    Mammalian Circadian Clock. PLoS Biol PLoS Biology.


    
              Karen Martin
GoldBio Marketing Coordinator


"To understand the universe is to understand math." My 8th grade
math teacher's quote meant nothing to me at the time. Then came
college, and the revelation that the adults in my past were right all
along. But since math feels less tangible, I fell for biology and have
found pure happiness behind my desk at GoldBio, learning, writing
and loving everything science. 



Category Code: 79102 88241

Posted by Karen on March 19th, 2015  ⟩  0 comments

The difference between growth factors and transcription factors might be a nonissue for many researchers, but after Google searching, I have discovered that the question does exist within our field.

My introduction to transcription factors came from my undergrad developmental biology class. The problem is that my professor classified both growth factors and transcription factors, all as transcription factors. And since this was my first time learning about it, there was no reason to question his lecture. I must also admit that my textbook either did not do a great job defining the difference, or I just wasn’t looking for it. For me, things like fibroblast growth factors all fell under transcription factors. I didn’t consider them to be synonymous; instead, I thought growth factors were simply a subcategory.

It wasn’t until coming to GoldBio that I began to question the possibility of a difference between the two. I’m not alone in this revelation, which is why I think it’s important to highlight the defining differences.

Transcription Factors: Transcription factors are molecules that can bind either directly or indirectly to a DNA sequence and regulate the transcription of a particular gene or set of genes. They function in concert with other proteins, either blocking or promoting RNA polymerase. Transcription factors also have at least one, but sometimes more, DNA binding domains that allows them to attach to specific sequences or to DNA near the gene being regulated.

Growth Factors: Growth factors are generally molecules that are secreted and interact with other molecules or specified receptors to influence cellular behavior, including cell differentiation, healing and cell proliferation.

Key Difference: The major difference is that transcription factors bind to DNA while growth factors do not. Instead of binding to DNA, growth factors interact with other cellular molecules.

The Relationship: Part of the natural confusion comes from the relationships that exist between the two. As you learn about different signaling pathways, the primary goal when you’re first introduced to the topic is to understand the interaction between molecules and proteins within a particular cascade. Once you wrap your head around that, it’s easy to overlook some of the physical and chemical differences that exist between each player.

In a given pathway, a growth factor is secreted by a cell. It then binds to its cell surface receptor, and that interaction catalyzes a chain of actions within the cell. Binding leads to signal transduction until finally a transcription factor is activated, or a cell’s growth behavior is influenced.

Naturally, that means that all the interactions must be especially fine-tuned for optimal performance. This is why we see horrible developmental disorders or cancers when a particular protein is mutated. Blocking part of a pathway can lead to a disruption in the whole process.

To further clarify it, here is a list of known mouse transcription factors and here is a list of GoldBio growth factors, which are used in a wide variety of developmental research and cancer research.

So now the confusion is dispelled. Perhaps you didn’t even know you were confused.


    
              Karen Martin
GoldBio Marketing Coordinator


"To understand the universe is to understand math." My 8th grade
math teacher's quote meant nothing to me at the time. Then came
college, and the revelation that the adults in my past were right all
along. But since math feels less tangible, I fell for biology and have
found pure happiness behind my desk at GoldBio, learning, writing
and loving everything science. 



Category Code: 88221 79108 88253

Posted by Chris on November 7th, 2013  ⟩  0 comments

Since the early 1990’s, there has been an explosion of research and discovery around the problem of preeclampsia and the correlated growth factor, VEGF. Preeclampsia has been a menace of pregnancies for ages and still affects anywhere from 2-10% of pregnancies world-wide. It’s a disease that strikes most often in first-time mothers, marked with high blood pressure and proteinuria, can be debilitating and can also lead to even more serious complications. But despite that, it is also a disease that is poorly diagnosed, or only properly diagnosed once the symptoms have been alleviated, i.e. the baby was delivered.

In the last 20 years, scientists have accumulated a massive amount of information about the Vascular Endothelial Growth Factor (VEGF) family and its role in preeclampsia. VEGF is a signal protein that is strongly involved in angiogenesis, building new blood vessels during embryonic development or after injury. It’s also very important in the implantation of the placenta to the uterine wall, wherein during preeclampsia, there is a shallow implantation, with reduced arteriole development and a subsequent immune response by the mother’s antigens. Research has shown that pregnancies resulting in preeclampsia show a significantly reduced level of VEGF, as well as its Placental Growth Factor (PlGF), an integral member of the VEGF family during pregnancy. They’ve also shown an increase in the soluble form of VEGFR1, called sVEGFR1 or sFlt-1. sVEGFR1 binds VEGF and prevents it from hooking up with the normal VEGFR1 and continuing its proper course. This results in a decrease in available VEGF which appears to be causative to preeclampsia.

There are a lot (and I mean a LOT) of papers dedicated to, or relating to, finding a marker protein or system to predict when preeclampsia will occur. In fact, there were so many duplications of the same work (I will continue to assume that they were all terribly unaware of previously published work), it was difficult to find a novel paper that went BEYOND simply reporting the problem and the symptoms. But I did! Allessandro Rolfo et al., from the University of Turin, took one step further back to see what was originally causing the sVEGFR1 to overexpress and cause the cascade of VEGF problems and the eventual preeclampsia.

Preeclampsia MSCsRolfo’s group found that mesenchymal stromal cells (MSCs) contribute to the pathophysiology of the preeclampsia condition through the irregular overproduction of a host of proimflammatory cytokines in PE placental mesenchymal stromal cells (PE-PDMSCs) as opposed to normal PDMSCs. Some of the elevated growth factors included IL6, IL8, TGFB2, LIF and TNF-alpha, just to name a few. Even more interesting was that they were able to take PE-PDMSC conditioned media and entice the same overproduction of those cytokines in normal explant systems! It’s a remarkable find, and if the results prove true, a definite boon to medical research. Pushing back to find the root cause of the problem is necessary to treating the disease and the next step for Rolfo’s group is to find out if the preeclampsia process can be reversed. Stay tuned. I'm sure there will be some amazing discoveries in the future!

 
 

Rolfo, A., Giuffrida, D., Nuzzo, A. M., Pierobon, D., Cardaropoli, S., Piccoli, E., Giovarelli, M. & Todros, T. (2013). Pro-Inflammatory Profile of Preeclamptic Placental Mesenchymal Stromal Cells: New Insights into the Etiopathogenesis of Preeclampsia. PloS one, 8(3), e59403.

Category Code: 79105 

 
 

PS. At Gold Bio, we’re always interested in finding things that interest our customers (and readers here at the Blog). If there’s a topic you’d like us to cover or an article that you think we should review, please email me at: cmenne@goldbio.com! We’d love to hear more from you!