biological stains + cell auras

My first bone soup!

I've made my bone soup with Ionat's help, along with the help of different biologists who I had to ask for chemicals and math assistance from: Dr Peter Mark, Research Fellow and Dr Stuart Hodgetts, Research Associate Professor, as well as one of Stuart's PhD students. They generously gave me the micro amounts of steroids (Dexamethasone) and Vitamin C (Ascorbic Acid) needed for the soup. The other bone soup ingredients include antibiotics, blood, sugar and a chemical that helps cells differentiate to osteoblasts. As well, the media (liquid) that is basically the bone soup stock, the DMEM, is formulated to keep the cell cultures healthy by preventing ammonia build-up. That build-up is like cell pee, to put it bluntly. As the cells eat and grow, they also produce ammonia, meaning they pee in their soup. They're kind of like pets in this way. Micro-organism lab pets, and in my case, bone pets.
I'll discover if the soup was properly mixed starting tomorrow when I extract bone cells from the marrow of a fresh bone (from the butcher) and attempt to isolate the osteoblasts. The bone soup is their supper and if they don't like it, I won't get healthy bone growth. Mixing the bone soup was *very* precarious. With some chemicals, the amounts were so small it was like a speck of dust, easily blown onto the floor with a misdirected breath. I had to weigh these chemicals on special, super sensitive scales that could measure in millionths of grams. So it is conceivable that I may not have made the best soup on my first go.

More calculations for bone soup recipe:

1mM Dexamethasone
10nM = 1 x 10[5]
1L (1000 mL) = 1,000,000 µL
so 500mL = 500,000 µL
5µL = 1 m....* in 500mL (*I can't read Peter Mark's writing here)

ß-GlyceroPhosphate (BGP):
for 50mL = 10.8mg for 1mM final concentration
for 500mL = .108mg for 1mM final concentrate
MW is 216.04

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When I checked on my Mighty Mouse cultures last Friday, I saw the aura around the cells! This edge of light around each living cell was vibrant and clear - I've never seen a cell aura before, and I have looked at C2C12s under a microscope previous to this. Maybe I didn't notice the aura before, but we'll just say that Amber did a good job with her energy transfer. The cell aura indicates that the cells are alive and healthy, their living light on display under the microscope. I was not using a photo imaging microscope so I don't have images yet, but I hope to soon.

***

In my work with bone decalcification, which started on Friday, I've moved from the chemical fixative stage (using Formalin, which is about 40% formaldehyde), to the de-cal stage (using Hydrochloric Acid), to the drying stage (in 70% Ethanol). This means that the fresh bones that I brought from the butcher (chicken wing bones) have already gone through three different chemical processes. They have about another nine chemical processes to go through yet this week before I can cut and use them. I'm working with Mary Lee, the Scientific Officer at CELLCentral lab and Shirley Chang, CELLCentral's Technician.

CELLCentral specializes in Histology, the science of tissue, and I am learning the old school craft of Histology from Mary and Shirley. The process of decalcification is to prepare bone for penetration by a wax support, so that the bone can be sliced at 5 microns thick, applied to glass microscope slides and dyed for viewing under the microscope. WHAT IS TRULY FASCINATING, for me and for all of my fibre arts/ textile friends, is learning that many of the tissue practices involve direct links to textile practices. For example, the dyes used in Histology originated from and are the still the same as textile dyes, including natural dyes like wood, saffron, coffee, indigo, cochineal, India ink, etc. Tissues which take up stains are called chromatic. Chromosomes were so named because of their ability to absorb a violet stain. Collagen sucks up stain really well. We are all colour and light, each of us. You can read more about staining HERE. Some examples of dyes/ stains at my disposal at the CELLCentral lab:

Vital Red Blood Volume, also known as fuchsine, is also a textile dye and is highly toxic. This will stain cell nuclei.

Canada Balsam is actually used as a cement for glass slides, because it dries optically clear. It is also edible.

Gentian Violet, also known as Crystal Violet, has medicinal properties as an antibacterial and antifungal. It isn't a textile dye, but it is used to dye paper, and is a nontoxic stain for DNA. It will also get rid of ringworm, candida and mouth ulcers. It has also been used to develop fingerprints in forensics (which as you may or may not know, is a passion of mine, being a bone woman and all). It's also used to mark skin for body piercings.

Carmine, a.k.a. COCHINEAL, is a natural textile dye made from beetle blood.

Sudan stains like fats, so are good for staining fatty tissue. This originates from a textile dye used to dye leather. It's also used for enhancing fingerprinting, since it likes body oils and grease.

Orcinol is a natural dye that comes from lichens. It is made with ammonia, meaning it used to be made with urine. It's also sold as cudbear paste and is used as a food dye as well. It will stain chromosomes and elastic fibres. It can also dye wool and silk without a mordant, anything from red to purple to blue.

This is also a textile dye used in bio staining. It's used to stain fatty tissue as well. It is toxic and promotes tumours and mutation. Do we want this rubbing off on our skin?

In histology, colour (dye/staining) is used as a diagnostic tool. It isn't simply to produce beautiful coloured images for the microscope, although they ARE stunningly beautiful. Different dyes adhere to different types of tissue. One dye type might show up cartilage primarily, while another might show up specific minerals or cells. This is fascinating to me. When I create my slices of bone that I then adhere (with wax) to glass slides, I can play with different dyes to see what emerges on each slide. Perhaps one slide will be dyed blue, the other with coffee. I'm also going to collect some eucalyptus leaves from the trees on my street and create a dye bath from that, which I will also use on my bone slides. I wonder what tissue type eucalyptus will stain!?


To give an actual colour example of what dyed/stained tissue looks like, I offer for your viewing these macro images I took today (I have a great macro lens on my iPhone) from already existing slides in the histology specimen library that I was able to pick through today and look at under the microscope.

WARNING: these are images of human fetal specimens.

Please keep in mind that they were thoughtfully and respectfully donated to science for the purposes of advancing human health.
They are absolutely stunning and were incredibly touching just to hold in my hand.
Please do not download or reproduce these images in any way. Thanks, dear friends.

Human fetal spine, length-wise.

Human fetal spine, cross-section.

Human fetal hand.

Human fetal foot.

The actual size of the specimens.

You can read up more on histology HERE.
This is a GORGEOUS old book in full downloadable format - my gift to you (click the text to download the beautiful PDF):
Conn, H. J. Biological Stains; A Handbook of the Nature and Uses of the Dyes Employed in the Biological Laboratory. Baltimore, MD: The Williams & Wilkins Company. 1953.