The Xanthophyll Cycle in Photosystem II

Wednesday, 18 June 2014

A Super Long Day: BBY Spinach Protocol

So today was an extremely long day in the lab. I was in from 8am-6.30pm...

The length of the BBY spinach protocol is why we started off with the isolation of stacked thylakoid preparation first, to see if it yielded high amounts of PSII super complexes. However, no matter what, the PSII super complexes were not pure in the sense that the PSI was still always attached onto them, which is not what we wanted. We only wanted PSII super complexes to analyse the pigments in the Xanthophyll cycle under light stress. Therefore, we took the the protocol of BBY spinach, which involves more steps, but the isolation of ONLY PSII super complexes is the final outcome- an outcome which we desire.

Due to the fact today was a  long day, I took many photos to keep me going & the 2 shots of coffee I took at 7am! (Warning you, there are many photos to follow, to the end of the page!)

Preparing the muslin filter ate for the grinded  slushy spinach leaves via the polytron. The protocol of BBY spinach starts off identical to isolation of stacked thylakoids.

Irina & Petra walking down the fourth floor corridor. Irina is a 2nd year biochemistry student from Queen Mary. She is doing her third year project with Ruban and basically is shadowing/ learning techniques before the start of her project! So glad she is here, more people in the lab more fun, as she is quite social so there's always something to talk about. Labs are always so fun in our team: Me, Irina, Petra and Erica. We have good chemistry, chemistry is so important. Can't imagine how my studentship would be like if it wasn't like this....

Fourth floor corridor, offices and labs ~

Walking up the stairs~

Irina university grown peas! Her 3rd year project will be involving peas so she has to grow them! It's quite nice how she can grow hers and see her material from start to end.  Due to the fact that my material is spinach and we buy from Whitechapel market, I have no control over the conditions the spinach are grown in! Obviously, every factor matters in science and we like to control as much as we can :)

Such a fine day on Fogg's rooftop ! Love the colour of the sky.

A fig tree grown by Anna, phd student at Ruban's Lab. Quite impressive right? Looks so healthy ;)

Adding the spinach leaves from the freezing cold icy water into the glass square cylindrical column, getting it ready for grinding with the polytron!

OH GOSH THE GREEN IS JUST SO LUSH! Love this photo, its ever so vivid ;) So yes filtering.

Squeezing the filtrate to get all the liquid out, you can only do this in the first filtrate as the second time filtering, you just let it filter drip.

#2 filtering 

Helping each other with the filtrate.

Adding solution in a filtering system. Later you will see how we filter the solution with pumping air outwards using a pump attached onto the de gaser equipment. Which rapidly forces the liquid to filter through the pores.

Measuring the O/D. Allowing us to calculate the chorophyll A: B ratio with such measurement. 

The blue slot is for the reference, acetone, what we added to our solutions extra.

Keypad for the spectrometer.

Pouring our solutions into the glass columns after centrifugation.

Nicely drawn table for recording the O/D of my solutions.

Moved labs where we shall be de gasing the solution so it will filter through rapidly. Petra is sorting out the pipes for vaccum out the air.

Me holding onto the pipe which vaccums out the air. As you can tell already the liquid on the top has rapidly filtered all the way through.
We have to do this knee tapping technique to get rid of the bubbles at the bottom of the flask. Alot of bubbles are attached to the magnetic stirrer which is required when we are making the buffer solution, adding all the powders.

Here is Ruban's dark lab! Instrumental !

Here is where all Ruban's researchers are placed. Erica, Petra, all the phd students: Anna & Max.

Biochemistry lab~ one which i've talked about already. I so prefer being in the biochemical lab there usually some form of light coming through compared to the dark lab. The dark lab just makes me feel sleepy during the day, as its pretty much blackout in there!

 Lunch time has arrived! So here is how my lunch is usually spent!

Finally I have manage to take a picture of the Fogg Building, which is Queen Mary's School of Biological and Chemical Sciences department. Yes completely lego, have to abit its one of the coolest buildings, or at least striking of that on campus ;) 

So we walked to the opposite side of the canal by the university to have lunch. What you observe: the blue building is student accommodation. Loving the hole in the middle of the building, in chinese feng shui terms its very good for a building to have a structured hole like that in the middle~ This is most of QM most expensive halls~ think its £180, but they get a bath tub ensuite! Posh! My first year halls is the brown halls right next to it on this right hand side of this photo. It's called Pooley house~ ensuite~ have fun memories there. Oh gosh how first year seems so ever long ago, but yet has passed so recently! 

By the canal, see this pretty canal boat with flowers on the top. Have to admit this area is a pretty nice area to relax and go for lunch on a hot summers day :)

My packed lunch bag. Yes it is abit childish, but I'm quite into all the cute things :) You are never too old, haha! Also, because I'm small and orential I can probably get away with it. Also, I brought a Costa (there is a Costa opposite my university) ice coffee to keep me going after lunch, especially because we have been in for so long, and had even more hours to do after ! 

Peak into my lunch box. Love siesta lunchboxes very clever designs! Healthy food = healthy mind :) good motto!

 Back to the labs!
Really love the labs timer. Its magnetic and fits nicely into my small hands. So need one for my oven as it doesn't have a timer, this piece of tech would be extremely useful!

This is a larger centrifuge, which can spin to max speeds of 30,000 rpm! Looks like a japanese washing machine to me :P

Because the large centrifuge doesn't have a pre cool setting, we keep the inner disc parts in the fridge to pre cool them that way!

Oh how UK weather changes so rapidly. Now it is gloomy! But the nice lego architecture of Fogg building always brightens any day!

This looks like it could be part of a canon! It is where we place out test tubes for centrifugation.

Massive microwave! Well looks like this, this moves whatever you put in it in a square motion!

Placing in the fridge already pre cool disc part of the centrifuge.

Fixing on the lid.

Seriously looks like a japanese washing machine...

Adjusting the settings.

Here is is in lab.

Ruban's office is very close to the large centrifuge, so I thought to take a photo :) Where all the meetings are!

Very concentrated material!

Me using a 5ml pipette, look so ready in action.

Inverting the tube to ensure uniformity in solution.

take #2 : smiles :D

Counter balance of water, checking via eye that it is roughly the same in level. Obviously, we measured it out before hand in weight to ensure it is correctly balanced.

Placing the tubes in!

Signing in that we used the centrifuge last. 

Using a paint brush and added H20 to re suspend the pellet obtained from the large centrifuge, important to keep in icy temperatures as mentioned before!

Using a syringe to filter through the material through a mini filter placed on top of the ependorf tube.

 End of the day: Autoclave time:

So there is a trolley outside the autoclave room which all the labs on the fourth floor place their equipment on, which they desired to be autoclaved.

Placing all the equipment in the trolley in the autoclave! 

Autoclave in a different position. Seriously looks like a massive pizza oven! I swear I always think that everything in the lab looks like something else in life, haha.

The instructions for which setting to set for what you are autoclaving. Pretty self explanatory.

Everything set. READY TO GO.

Finally, the end of the day: What a long a day! 
And also a blog post too! Haha hope you enjoyed it :)

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