Étiquette : understanding experiments

Everyday Objects in Science

Le par julienforsciencedans Understanding Science / Comprendre la scienceLaisser un commentaire

Today’s article will be a bit different. The idea came to me as I was working in my own lab, and I realized that we use a lot of everyday objects in labs, but not for the reason they were invented. So today, I will give you five example of everyday objects that scientists use for their experiments.

Paintbrush: the unusual transport system

In a lot of labs, we perform an experiment called immunohistochemistry (IHC), which allows to see tissues under the microscope, and use antibodies to label specific proteins. For this experiment, the tissues are so thin that they will break very easily. That’s why, instead of a spoon or any other utensil, we use a paintbrush to move the tissues around. Since it is very soft, it won’t break the tissue. Further, a paintbrush is somewhat bendy, which allows for easy scoop up of tissues.

Nylon Stockings: how to keep the tissues in place

This one might be specific to only a few labs, as it is useful for a technique called electrophysiology. This technique allows us to record the electric currents in brain tissues. To keep these tissues alive, we keep them in an artificial liquid, called artificial cerebrospinal fluid. Since it is in a liquid, it has the tendency to move around a lot. To prevent this, we create a special tissue holder, made up of threads from a stocking. These thread are thin but strong, thus they won’t break but will keep the tissue in place.

Powdered Milk: the blocking agent

Milk is a staple element in most biochemical labs. Many experiments use antibodies to label proteins, but unfortunately they have the potential to bind to another protein that is not the one you want to label. This is called unspecific binding. To prevent this, we need to block our tissue. We use proteins that will bind to any unspecific binding sites, thus blocking them. The antibodies can then be placed on the tissue with no worries. Milk contains a protein called casein which can do just that [source].

Aluminum Foil: the tissue’s umbrella

When we use antibodies, oftentimes they are coupled with a fluorescent tag, which allows us to see the antibody, and the protein it binds to, under a microscope. However, these fluorescent tags are very sensitive to light. If exposed for too long, it will not be able to fluoresce anymore, because the photons in the light damage the tag. This is called photobleaching. To prevent this, we put our antibodies in the dark. However, when we want to move the antibodies around, we use aluminum foil. Aluminum foil does not let light pass through and thus protect any antibodies from photobleaching [source].

Nail polish: an unlikely glue

Again, this is used in IHC experiments. In these experiments, the tissues are mounted on a glass slides, and covered with another glass coverslip to prevent movement. However, instead of glue, we use nail polish to glue both glass pieces together. we use it mainly because we can apply it more easily than glue, and we know that it will not interfere with the fluorescence. Further, if clear nail polish is used, we will be able to see through it under the microscope.

There are many other everyday objects that scientists use. From microwaves to coffee filters, a lab is surprisingly made up of things everyone knows. This shows how sometimes science is more accessible than we think it is.

Understanding Science: In Vitro models

Le par julienforsciencedans Understanding Science / Comprendre la scienceLaisser un commentaire

Today’s article is a continuation of the last one. We first looked into the model organisms, or in vivo models, and their advantages. However, it is very hard to use in vivo models, due to the strict ethic codes associated with them. Therefore scientists developed ways to perform experiments without the use of animal models. These models are called in vitro models (that can be translated in model in glass). The principle of an in vitro model is simple: isolate the component of interest, and study it away from its source. This article will present different in vitro models and their advantages over in vivo models.

Polymerase chain reaction: a way to get « infinite » DNA

The first example is a staple of any scientific lab. Polymerase chain reaction (PCR for short) is a technique invented by Dr. Kary Mullis (who got the Nobel Prize for this finding), and it allows to artificially replicate DNA. The technique works very similarly to our own cells. In our body, we have proteins called helicase which separates the DNA to allow another protein called polymerase to bind the DNA and replicate it. This is however very energy intensive and hard to replicate in vitro. PCR uses an easier way to separate DNA: temperature. Under high heat, the DNA will separate by itself and allow the polymerase to bind. Therefore, when performing PCR, we add the DNA we want to replicate, and polymerases. The DNA will undergo many heat cycles, allowing it to separate, and the polymerase will then replicate the DNA. With another cycle, the newly replicated DNA can be separated, and more DNA will be formed. Hence, we get exponentially more DNA than what we had in the first place.

This technique has many applications. In the medical field for example, simply taking the DNA of a patient from the saliva can allow us to replicate the DNA indefinitely and study it to know which genes are mutated for instance. But more interestingly, we can replicate DNA which we have limited access to in order to study it indefinitely. That’s how we know the genetics of some dinosaurs. The limited DNA preserved in fossils was replicated using PCR, and then studied more extensively without the risk of losing the DNA forever. Finally, hair strands or any other organic materials found in crime scenes can be used in PCR to study the genetic fingerprints and find the culprits. PCR is an amazing technique that helped us preserve DNA we would have lost, and because so much DNA is produced, we can use it indefinitely for many studies [source / source / source].

Protein purification: isolating a protein to study it by itself

Similarly to DNA, we can also isolate proteins. However, unlike PCR, we will not replicate it indefinitely. Instead, we will separate it from everything else to study its structure, or see what its effects are. Since there are so many proteins that are doing a lot of things at the same time. It is hard to determine what a single protein is responsible for, That’s why isolating it makes it easier to study. There are many ways to separate proteins, some of which I previously explained, such as Western blots or flow cytometry. I wanted to mention a last one, and maybe the most famous, centrifugation. Centrifugation allows to separate particles depending on both density and speed. When a sample is centrifuged, any molecule that is affected by the speed will aggregate at the bottom of the tube, while the other will stay suspended. In the cases of proteins, it is often used to separate all the proteins from any other components of the cells. In short, centrifugation is a crude way of separating different components within a sample [source / source].

Cell culture: getting cells to do what you want

Cell cultures are probably the most common in vitro models. As the name suggest, we are taking cells from a organisms, and culture them in Petri dishes. One advantage of cell cultures is that they can become immortal. using a process called transformation, we make the cells express genes that prolong their lifespan to the point that they will never die. Further, we can keep cell cultures forever by simply freezing them, preventing any unwanted reactions since cold proteins will not activate. Cell cultures have a lot of advantages. First, we can perform invasive or « painful » experiments on them, since it is outside of the organisms and thus no pain will be felt. We can also culture human cells, allowing us to understand some diseases better than if we were to use animal models. Lastly, simply from skin cells, we are able to make any cell we want. This procedure is called induced pluripotent stem cells (or IPSC), and it allows us to revert any cell into a stem cell. A stem cell has the capacity to become any cell, therefore we can transform the skin cells of a patient into neurons to better study them without having invasive surgery [source / source / source / source].

Here we have seen three example of in vitro models and how useful they can be. In many instances, they have advantages over in vivo models. First we can study human tissues, but we can perform more painful or dangerous experiment without any risk. However, in vitro models have their flaws. The first one is that they still depend on biological sample. Every example that I have given still requires an initial sample from a patient or an animal. This implies that we still need ethical conducts towards in vitro models, notably how and how long we are going to use the sample, and who will have access to it. Lastly, the core principle of in vitro models is isolation, and it is its biggest flaw. The human body is so interconnected that all of its parts influence each other tremendously. When we isolate something from the body, it will not react the same way. This is why many times results found in vitro will never happen in vivo. This means that many in vitro experiments have to be repeated in vivo in order to be sure that the results we see is actually happening in the body.

Understanding Science: model organisms

Le par julienforsciencedans Understanding Science / Comprendre la scienceLaisser un commentaire

Our article today will again aim to understand how we do science. This time, we will study what is named model organisms. These are simply living organisms we use to achieve our studies. Now I know this topic is highly controversial. Therefore, I hope that this article helps to show how essential studies in living organisms, also called in vivo studies, and how much they’ve contributed to overall science. Next week, we will study models that does not involve animal, or in vitro studies, and their differences.

The first question to answer is why are we doing in vivo studies? The simple answer is because we still don’t have any other choice. While science has advanced tremendously, and as we will see next week we have a lot of choice when it comes to study model, none will provide what an animal can. The most obvious reason is drug testing. Let’s say a lab has found a potential drug to treat heart disease. After years of studies without any animal, the lab can confidently say that it works in vitro. However, that does not mean that it will in vivo. Most in vitro models do not have a very long lifespan. What if this drug increases the chances of cancer? The best way of knowing this is testing it on an animal, and see potential side effects. Now, it is illegal to try any medication on animal if no previous intensive studies have been made. We will see later the ethics of animal studies, but one of the staples is animal comfort, thus injecting drugs with no idea of what they may do is not only illegal but also punishable by law. In conclusion, we use animals in this setting to make sure that the drug is safe in humans. But we also use animals to study diseases. None of our in vitro models will be as accurate as in vivo model of diseases. Thanks to these animal, we found countless medications and saved many lives in the process. Now we will go through examples of in vivo models and their uses throughout science [source / source / source / source].

Yeasts and Fruit flies: staples of genetic studies

We start with rather unusual yet essential model organisms: the yeast, specifically the one called Saccharomyces cervisiae. This species was essential to understand how genes and genetic overall works. As a small organisms, its genome (meaning the entirety of their genes) was mapped and understood very quickly. This in turn helped us understand how genes interact with each other, and how their transcription and translation works. Furthermore, yeasts reproduce extremely fast, which allows us to study genetics and development over multiple generations faster than any other organisms. The same can be said fruit flies, specifically Drosophila melanogaster. Fruit flies have the same advantages as yeasts, being fast breeder, cheap, and easy to genetically map, with the added bonus of being anatomically diverse. While the yeast only looks like a blob of cells, flies have distinct features. This is essential because they allow us to understand the relationship between genetic and development. For instance, we can know which gene allows the production of legs, and which one causes eye color. However, both of these organisms also have disadvantages. The main one being that they are genetically quite different from humans. Although we found genes in yeast and flies that were similar or identical in humans, many discoveries are hard to relate to humans. Furthermore, these organisms lack quantifiable behaviours, making it hard for us to link genetics and behaviours [source / source / source].

Frogs: understanding proteins and cells.

Still unusual, a specific type of frog offers unique tools to study the behaviours of cells and proteins. The Xenopus laevis is an African frog that has very interesting eggs, or oocytes. Indeed, one frog makes a huge amount of eggs, with an interesting particularity: each cell in it has a pre-determined role. It means that we know exactly what each cell will do to become a tadpole. This is extremely interesting to study development and cell behaviour. We can know, for instance, how the morphology of a tadpole will change if we slightly move a cell in the oocyte. The usefulness of these oocytes is enhanced by the sturdiness of these cells. Xenopus oocytes can withstand a lot, from injection to rough manipulation, and still yield a viable tadpole. It allows us to inject genes or protein and see what they do to the cell behaviour or the development. Lastly, eggs and be cultured and used for in vitro studies later. But what’s interesting with oocytes is that they allow us to inject several proteins and see how they interact with each other. This allows us to better understand protein behaviour as well [source / source].

Mammals: understanding diseases and complex behaviours

Mammals are amongst the most used animal models in science, and for a good reason. They have complex behaviours and thinking processes, easily comparable to humans. We use many different mammals, but the most common ones are mice, rats, and primates. Studying them as is is an invaluable tool to understand behaviour and relate it to humans. But more interestingly, we can study behaviours and physiology in context of diseases. While similar, there are big differences between rats and mice which will influence your research. Rats are bigger, easier to manipulate, and closer to humans in terms of genetics and behaviours. But they have a big disadvantage: it is hard to genetically manipulate them. Mice on the other hands are very easily manipulated genetically. This allows us to have mice models for many diseases, and we can study the disease and see its effect on our cells. Primates are great to study group behaviours. As they are closest to us, they can tell a lot on how cognition, memory, and complex behaviours work. However it is extremely hard to work with primate, and it’s almost impossible to genetically modify them, although very recently it was done [source / source / source / source].

The ethics of animal care: the 3Rs

Working with animals in Canada is not only difficult but also very regulated, and rightfully so. The Canadian Council on Animal Care (CCAC) set up many rules to follow when scientists use animals, and the basis is known as the 3Rs. First, Replacement forces us to prove why we need animal. Animal studies are last resort, and unless you prove that no in vitro model can do what you want, you will be denied animals. Second, Reduction forces us to optimize animal use. You have to determine how many animals you need per year, and not only does it have to be the smallest number possible, to have more animals than requested is very hard. Lastly, Refinement forces us to place animal comfort above all. No animal should suffer or be stressed for useless reason. Any amount of pain or stress has to be proven useful for the experiment, and animal comfort is ensured while the animal is not used. For example, mice are not kept separately but rather in group (unless the experiment asks for single housing). Similarly, primates have mandatory physical, social, and play time every day. For any more specifics on animal care, the CCAC website is very thorough [source / source / source / source].

In conclusion, I hope that I managed to show how important animal models are for science. I understand how uncomfortable the topic is, but it is important for the public to understand that not only we are treating animals the best we can, but that we have no other choice, as right now they constitute the best model we have to help human society.