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Differential

centrifugation

Muscle tissue HomogenisationTissue homogenate

Contractile apparatus

Mitochondria

60 min at 100 000g

Crude

microsomes

Cytosol

Gradient

centrifugation

Homogenisation

10–60% Sucrose density gradient

360 min at 150 000 g

Surface membranes

Triads

Light sarcoplasmic reticulum fraction

Heavy sarcoplasmic reticulum fraction

Debris

10 min at 1000 g

10 min at 10 000 g

20 min at 20 000 g

SupernatantNuclei, cell debris

Extracellular matrix

Sarcolemma

(b)

(a)

Supernatant

Supernatant

Cytosol

Longitudinal

tubules

Non-

junctional

transverse

tubules Triad junction

Mitochondria Skeletal muscle fibre

Terminal

cisternae

Sarcoplasmic reticulum

Scheme of subcellular fractionation of membranes from muscle homogenates

Subcellular membrane systems that can be isolated by differential centrifugation

Fig. 3.5Scheme of the fractionation of skeletal muscle homogenate into various subcellular fractions.

Shown is a diagrammatic presentation of the subcellular membrane system from skeletal muscle fibres

(a) and a flow chart of the fractionation protocol of these membranes from tissue homogenates using

differential centrifugation and density gradient methodology (b).

90 Centrifugation