Protokol Penyediaan Sampel utk TEM ( Transmission Electron Microscopy )

1.Primary fiksasi
-2.5 % glutaraldehyde dgn 0.1M buffer / 1-2% glutarldehyde + 1-4% formaldehyde selama 4
jam.

* proses fiksasi formaldehyde lagi cepat fiksasi berbanding dgn glutarldehyde.ini utk mengelak
tissue mengecut terlau cepat dan mengelak drp tissue itu dianggap abnormal, padahal tisu itu
normal.

2.Basuhan/bilasan
phosphate buffer/cacodylate selama 30min, 3 kali

3.Secondry fiksasi
1-4% osmium terroxide dgn buffer selama 1-2 jam shj.pada peringkat ini masa tidak melebihi 2
jam, ini kerana jika terlalu lama struktur tissue akan menjadi terlalu hitam.

4.Basuhan/bilasan
phosphate buffer/cacodylate selama 30min, 3 kali

5. Tertiary fiksasi 1-2% uranyl acetate selama 30min

6.Basuhan/ bilasan
buffer/ air suling 30 min, 3 kali

7.Proses Penyahidratan ( Gaya UKM ) dia guna alkohol

-50% 15min
-70% 15min
-80% 15min
-85% 15min
-90% 15min
-95% 15min
-absolute ethanol / isoamyl alcohol 30min diulang 2kali

*Kepekatan sesuatu alkohol bergantung kepada jenis sesuatu sampel berkenaan, akohol dan aseton boleh digunakan sebagai ajen penghidratan, walabagaimanapun aseton lebih ekstrem dalam melakukan proses dehidrasi berbanding dengan aklohol.walauapapun, kedua-dua nya berfungsi sebagai bahan yg mengeluarkan air dlm sampel biologi yg hendak disediakan.

8.Embedding / resin / polymer

-1 % resin / 3 % aseton
-3 % resin / 1 % aseton
-100 % resin

*100% resin dalam kapsul conical BEEM ( tiub berpenutup )
*diperingkat ini, resin tidak akan menyebabkan proses polimerisasi jika sampel tidak di
dehidrasi dengan sempurna.


9.Polymerization
-8 jam /semalaman.

10.Ultrathin sectioning/ pemotongan seksyen ultra-halus
-ketebalan < 100nm

12.Kontras / Pewarnaan
-Pewarnaan Negatif
-Pewarnaan Positif

PCR song...its great n funny LOL

The PCR Song

There was a time when to amplify DNA,
You had to grow tons and tons of tiny cells.
Then along came a guy named Dr. Kary Mullis,
Said you can amplify in vitro just as well.
Just mix your template with a buffer and some primers,
Nucleotides and polymerases, too.
Denaturing, annealing, and extending.
Well it’s amazing what heating and cooling and heating will do.
PCR, when you need to detect mutations.
PCR, when you need to recombine.
PCR, when you need to find out who the daddy is.
PCR, when you need to solve a crime.
(repeat chorus)

Anyone who are involves in PCR in the lab sure they are the smile first :-) with this song huhu!

Enjoycess..! release ur tension b4 sit ur exam n viva test.

Refer Youtube at the right side..