Electron Microscope Imagery: “Virus” Proof or Refutation?

One of the “proofs” people try to offer in support of “SARS-COV-2” or any other “virus” is that pictures of these particles exist. They assume that since we can see images of these “viruses,” this means that they must be real. The images referred to are normally Transmission Electron Microscope images taken from cell cultures. However, there are numerous issues in relying on cell cultures for proof of anything as detailed here:

The Case Against Cell Cultures

Before the invention of the Electron Microscope in 1931, “viruses” were just assumed to exist and cause disease. They could not be seen visually. After the invention of EM, normally invisible particles claimed to be “viruses” could now be seen. However, the status of these particles as a “virus” was still assumed as they were never properly purified/isolated from an unaltered sample from a sick patient nor proven pathogenic in a realistic way using animal models. What virologists do is look for particles fitting the image they want of a “virus” in an unpurified cell culture sample (which contains potentially millions of similar-looking particles) and then imply pathogenicity as well as function to the ones they choose. Yet without purifying and isolating the particles assumed to be “viruses” first, there is absolutely no way any virologist could ever know that the particles they select as their “virus” truly are one.

This is a brief description of how the cell culture samples are prepared for imaging and the many particles that are certain to be in the sample which resemble “viruses:”

General Procedures for Electron
Microscopy Applications in
Diagnostic Virology

“Briefly, a 10 µl preparation is taken from the cell culture, placed on a formvar and carbon coated grid, this
is followed by the addition of 10 µl of negative stain (e.g. phosphotungstic acid). The solutions are left on the grid for a few seconds to 1 minute followed by drying with filter paper. The grid is then ready to be viewed using a TEM. With this method the background is stained and particles, including intact
virions are left unstained, therefore outer details of the virus are visualized against the electron-dense background. Care must be taken with interpretation since the sample contains other cellular debris that can be in the size range of a virus. Notice the cellular debris in the negative staining sample in Figure 1a;
careful interpretation must distinguish between virus and other cellular elements, such as broken down cellular membranes.”

“Ultrastructural interpretation is a critical component for the correct identification of viruses within a sample. When TEM techniques are applied to diagnostic samples it is often for the purpose of detecting a virus and thus there may be a bias to closely scrutinize the tissue to find a particle that resembles a virion. Generally in cell culture the virus has been amplified to such high concentrations that detecting and identifying the virus should be relatively straightforward. If it is not, then other causes for CPE in the cell line, such as toxicity, should be investigated. In cases when fish tissues are examined the viral concentrations may be lower. Strict criteria must be utilized in order to be confident that a virus is responsible for the associated pathology and caution must be practiced to avoid misidentifying normal cell structures that may resemble a virus. When a viral etiology is suspected and searched for in tissue it is amazing how many cell structures can resemble a virus!”

Host cellular organelles can fall into the same size range as viruses and can resemble viral structures, although clear differences discern cell organelles and virions. The potential for the structures in question to be related to cellular organelles in either normal or pathological states must be ruled out to avoid misidentification of cellular structures as “virus-like”. Several cellular components in the cytoplasm that may be confused for viruses include primary lysosomes, secretory granules (Figure 6a,b), transport vesicles (Figure 6c), glycogen (Figure 7), and crystalline inclusions. In the nucleus structures such as nuclear pores and perichromatin granules (Figure 8), measuring 30-35 nm, are very common and must not be confused with viruses found in the nucleus, such as herpesvirus and adenoviruses. Nuclear pores are found within the nuclear membranes and when sectioned en face the pores are clearly visible. In addition to normal cell structures, pathological processes can lead to unusual cytoplasmic or nuclear inclusions that can resemble viral structures. For some families of viruses, assembly of the virions is
associated with inclusion bodies that are composed of viral structural proteins, membranes, and sometimes mature viral particles. The inclusions may be single or arranged in a lattice structure, but should only be considered viral if mature virions are observed, since crystalline inclusions and tubular inclusions can occur within host cells without viral involvement. Figure 9 shows a cell with tubular inclusion bodies in the spleen of an apparently healthy rainbow trout. Though these inclusions are suspicious, no viral particles were apparent. Inclusions such as these can be a result of cellular degeneration. Other pathological processes including aging, toxicity, and misfolded proteins can lead to inclusion bodies involving host cell proteins (Cheville 2009).”

https://www.google.com/url?sa=t&source=web&rct=j&url=https://units.fisheries.org/fhs/wp-content/uploads/sites/30/2017/08/2.1.2-Electron-Microscopy-for-Virology.pdf&ved=2ahUKEwi3poS69pnvAhVJXK0KHUC7CT44FBAWMAB6BAgEEAI&usg=AOvVaw1iLCKhWcDiQKWNMtbIxVxi

The fact that there are many micro and nanoparticles within the culture which resemble “viruses” shows why it is absolutely essential for the sample said to contain a “virus” to be PURIFIED (i.e. free of any contaminants, foreign material, pollutants, etc) so that a “virus” is ISOLATED (i.e. separated) from everything else which could resemble one. Exosomes, which are identical to “viruses,” are some of the millions of “virus-like” particles that are guaranteed to be within samples:

“Viruses” or Exosomes?

TEM images of exosomes in Rabies Vero cell culture.

Besides the fact that there is no realistic way for a virologist to pick out a particle from an unpurified sample in a TEM image and claim it is the “virus” they were looking for, there are other disadvantages to TEM for “viral” identification as well:

Disadvantages of electron microscopy

“However, there are several disadvantages which may mean that other techniques, especially light microscopy and super-resolution microscopy, are more advantageous to the researcher. These include:

  • Inability to analyze live specimens – As electrons are easily scattered by other molecules in the air, samples must be analyzed in a vacuum. This means that live specimens cannot be studied by this technique. This means that biological interactions cannot be properly observed, which limits the applications of electron microscopy in biological research.
  • Black and white images – Only black and white images can be produced by an electron microscope. Images must be falsely colorized.
  • Artefacts – These may be present in the image produced. Artefacts are left over from sample preparation and require specialized knowledge of sample preparation techniques to avoid.

https://www.news-medical.net/amp/life-sciences/Advantages-and-Disadvantages-of-Electron-Microscopy.aspx

As the electron microscope kills anything subjected to it, there is no way to visualize living tissues/samples. The images are only able to be seen in black-and-white which is why we see so many digitally colorized and enhanced photos to sell us on the idea of “viruses.” Artefacts are almost a given to be present in some capacity due to the many alterations the sample goes through in order to be imaged correctly. There is no possible way that virologists can claim what is visualized in the EM images was ever actually in that state prior to being fixed and embedded for imaging. The process of fixing and staining a sample for viewing in a TEM image is guaranteed to alter the sample and carries several disadvantages:

Preparing samples for the electron microscope

“Electron microscopes are very powerful tools for visualising biological samples. They enable scientists to view cells, tissues and small organisms in very great detail. However, these biological samples can’t be viewed on electron microscopes whilst alive. Instead, the samples must undergo complex preparation steps to help them withstand the environment inside the microscope. The preparation process kills the tissue and can also cause changes in the sample’s appearance.

For scientists who wish to view biological samples, this poses a challenge – how can the sample be preserved so that it looks as much as possible like it would in the living organism, while still being able to withstand being visualised in the electron microscope.”

To be visualised by an electron microscope, biological samples need to be:

  • fixed (stabilised) so the electron beam doesn’t destroy them
  • dried thoroughly so the vacuum doesn’t affect them.

Fixation: a snapshot of the living sample

The first – and perhaps most important – step in the preparation process is fixation. In this step, living tissue is chemically treated to stabilise it. This kills the tissue sample at the same time. It’s important to fix a sample as quickly as possible because, as soon as tissue is removed from its natural environment, it starts to change. For instance, oxygen levels start to drop as soon as tissue is removed from an organism. This causes mitochondria to start to change their appearance. Another common change in the fixation process is that lipids tend to form micelles.

Looking out for artefacts of fixation

Micelles and strange-shaped mitochondria are examples of artefacts – structures that are seen under the microscope but aren’t found in living cells. It’s very important to be aware that artefacts can be introduced during fixation so that you don’t mistake them for real parts of your sample. Telling the difference between an artefact and a ‘real’ structure can be difficult.”

“For TEM, samples must be cut into very thin cross-sections. This is to allow electrons to pass right through the sample. After being fixed and dehydrated, samples are embedded in hard resin to make them easier to cut. Then, an instrument called an ultramicrotome cuts the samples into ultra-thin slices (100 nm or thinner). TEM samples are also treated with heavy metals to increase the level of contrast in the final image. The parts of the sample that interact strongly with the metals show up as darker areas.”

https://www.sciencelearn.org.nz/resources/500-preparing-samples-for-the-electron-microscope

Artefacts caused by dehydration and epoxy embedding

In Summary:

  • Care must be taken with interpretation since the sample contains other cellular debris that can be in the size range of a “virus” (i.e. cellular elements, such as broken down cellular membranes)
  • There is an admitted bias to closely scrutinize the tissue to find a particle that resembles a “virion”
  • If evidence of a “virus” is not found, then other causes for CPE in the cell line, such as toxicity, should be investigated
  • Caution must be practiced to avoid misidentifying normal cell structures that may resemble a “virus”
  • It is admitted that it is amazing how many cell structures can resemble a “virus”
  • Host cellular organelles can fall into the same size range as “viruses” and can resemble “viral” structures
  • The potential for the structures in question to be related to cellular organelles in either normal or pathological states must be ruled out to avoid misidentification of cellular structures as “virus-like”
  • Several cellular components in the cytoplasm that may be confused for “viruses” include:
    1. Primary lysosomes
    2. Secretory granules
    3. Transport vesicles
    4. Glycogen
    5. Crystalline inclusions
  • Nuclear pores and perichromatin granules are very common and must not be confused with “viruses” found in the nucleus, such as herpesvirus and adenoviruses
  • Pathological processes can lead to unusual cytoplasmic or nuclear inclusions that can resemble “viral” structures
  • Inclusion bodies (aggregates of particles) should only be considered “viral” if “mature virions” are observed, since crystalline inclusions and tubular inclusions can occur within host cells without “viral” involvement
  • Inclusions can be a result of:
    1. Cellular degeneration
    2. Pathological processes
    3. Aging
    4. Toxicity
    5. Misfolded proteins
  • There are several disadvantages to Electron Microscopy such as:
    1. Inability to view samples alive
    2. Black-and-white images only
    3. Production of artefacts which can be mistaken for “viruses”
  • Samples must undergo complex preparation steps to help them withstand the environment inside the microscope
  • The preparation process kills the tissue and can also cause changes in the sample’s appearance
  • How can the sample be preserved so that it looks as much as possible like it would in the living organism, while still being able to withstand being visualised in the electron microscope?
  • Samples are chemically treated which kills them before imaging
  • This process must be done quickly because as soon as the tissue is removed from its natural environment, it starts to change
  • Mitochondria will start to change their appearance
  • Artefacts can be introduced during fixation and telling the difference between an artefact and a ‘real’ structure can be difficult
  • After being fixed and dehydrated, samples are embedded in hard resin to make them easier to cut
  • TEM samples are also treated with heavy metals to increase the level of contrast in the final image

There are many chemicals used and procedures done to the cell culture sample before the imaging can take place. It is admitted that these processes can alter the sample which introduces artefacts. These artefacts are structures not seen in living cells yet the sample that is seen in an TEM image is no longer living and has been heavily altered not only by cell culture conditions but also by the fixing and staining process. For a virologist to claim any of these particles are “viruses” let alone that they are found in living cells or hosts is disingenuous at best and flat out lying at worst.

Look at the TEM images below. Can you tell which are “viruses” and which are exosomes or other extracellular vesicles which resemble “viruses?”

Virologists can not either.

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