Section D

Live Dye Partial Organells and Observe Them with Microscope

April,2013

Department of Cell Biology School of Basic Medical Sciences

Xinjiang Medical University

Nafeisha Kadeer

Purposes:

• Master the method to live dye partial organells--mitochondria and vacuole system.

• Master the basic structure and distribution of mitochondria, vacuole system in cell.

Contents:

1.Live dye the mitochondria in the rabbit liver cell.

2.Live dye the vacuole system in the swordlike cartilage cell of toad.

3.Observe the Golgi Apparatus.

Materials

1. microscope,slide,coverslip,pipette, tweezers,absorption tissue,dissection

plate,dissection needle,dissection scissors,eye scissors,culture vessel,nails,thick plastic gloves;

2. a rabbit,a toad;3. 1/300Jaun’s Green 1/3000Neutral red 0.85% Ringer solution 0.65% Ringer solution

1 、 The method to live dye the mitochondria in the rabbit liver cells

Principle

Jaun’s Green is a special live dye for mt. The

cytochrome oxidase in the mt can oxidize this

dye into blue particles, so we can examine the

existence of the mt by these blue particles.

Inject air into rabbit’s ear vein

vein

artery

Method • Kill a rabbit (inject air into it’s ear vein) , put it on

the dissection plate on back, then open it’s abdominal

cavity. Cut down a small piece of liver tissue from thinnest rim, put it into cultured dish, with Ringer’s solution wash off the blood.

• Transfer liver tissue on slide, make it vertial. From it’s bottom add one drop of Jaun’s Green to half-cover it, stained for about 20 minutes.

• Dispart tissue using tweezer, remove the large blocks tissue. Finally some single cells are left on slide.

• Add a drop of Ringer’s solution, then cover it with coverslip, absorb excess solution.

• Observe your sample with microscope.

Observe the slide in the microscope.

Using the objective of 10x magnification, then

the objective of 40x magnification.

We can see there are many blue particles in the

cytoplasm of the liver cells, which are arranged

tightly. These blue particles show location of mt.

Result:

Mitochondria in liver cell

nucleus

cell membrane

mitochondria

(high power objective)

2 、 How to live dye the vacuole system in the swordlike cartilage cells of toad

Principle

In cytoplasm of animal cell, the vesicles enclosed by the

membrane all belong to vacuole system (except mt), including

Golgi Apparatus 、 Lysosome 、 Endoplasmic

Reticulum ， etc. The cartilage cells contain abundant vacuole

system. However, these vacuoles are too small to be seen if they

are not stained with dye. Neutral red can specially stain the

vacuole system dark red and stain cytoplasm and nucleus light

red. So we can observe location of vacuole system .

Method • Kill a toad, open it’s abdominal cavity, then expose

the swordlike cartilage of breastbone.

Cut down a small piece of cartilage tissue from its thinnest rim, put it on slide.

• Add a drop of Neutral red to dye for 15 minutes (not more than 15 min).

• Remove the dye using the absorbent tissue. Add a

drop of Ringer’s solution, add coverslip.Absorb

excess solution.

• Observe your sample with microscope.

Swordlike cartilage of toad

swordlike cartilage

Under microscope, we see cartilage cell is

oval. There are many rosy vesicles .These

rosy vesicles are the vacuole system.

Result:

3 、 Observe Golgi Apparatus

There are prepared slide of rabbit nerve

knot. First observe it using low power. We can

see many oval neurons. It is oval and stained yellow. Then observe

the neurons using high power.

We can see the round and transparent central parts are nucleus of

neuron. In cytoplasm there are many dotlike 、 sticklike and

threadlike structures scattered and stained dark tan. These dark tan

structures are Golgi Apparatus.

(high power objective)

nucleus

nucleolus

Golgi Apparatus