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Organ Culture
Animals - 16-1S-day-old oothecae of the American cockroach Periplaneta
amencana
• • Procedure
Sterilization 1. Surface sterilize the oothecae by submersion in 70% ethanol for 5 min.
2. Wash the eggs in sterile distilled water.
Dissection 1. Remove the embryos from the eggs in culture medium placed in the
depression of a Maximov slide.
2. Excise the brains, sub esophageal ganglia, thoracic ganglia, and the entire digestive tract.
Culture 1. Place the excised organs onto a small cover glass (10 x 10 mm).
set-up 2. Place the cover glass into a culture vessel containing O.IS ml culture
medium.
3. Place five or six culture vessels in a petri dish, and store them in a
desiccator containing 95% air and 5% CO2•
4. Incubate the culture at 29°C in a water-saturated atmosphere .
• • Results
According to Chen and Levi-Montalcini (1969, 1970), the brains or ganglia showed axonal outgrowth by the end of the 2nd day of culture. The
outgrowing axons branched repeatedly, and formed connections with
each other and with the outgrowth of other explants. A small enlargeFig. 3. Photomicrograph (Nomarski) of living 3-month-old culture of dissociated
nerve cells, thoracic ganglia (th), and foregut (j) explants from 16-day cockroach
embryos, Periptaneta americana. Nerve fibers branch out in large number from the
ganglion and interconnect with fibers produced by dissociated nerve cells. x 20.
From Levi-Montalcini and Aloe, "Neuronal nets and nerve cell interactions in vitro
in insect systems:' In Vitro 8, pp184. Copyright © 1972 by the Society for In Vitro
Biology (formerly the Tissue Culture Association). Reproduced with permission
of the copyright owner
Organ Culture
Animals - 16-1S-day-old oothecae of the American cockroach Periplaneta
amencana
• • Procedure
Sterilization 1. Surface sterilize the oothecae by submersion in 70% ethanol for 5 min.
2. Wash the eggs in sterile distilled water.
Dissection 1. Remove the embryos from the eggs in culture medium placed in the
depression of a Maximov slide.
2. Excise the brains, sub esophageal ganglia, thoracic ganglia, and the entire digestive tract.
Culture 1. Place the excised organs onto a small cover glass (10 x 10 mm).
set-up 2. Place the cover glass into a culture vessel containing O.IS ml culture
medium.
3. Place five or six culture vessels in a petri dish, and store them in a
desiccator containing 95% air and 5% CO2•
4. Incubate the culture at 29°C in a water-saturated atmosphere .
• • Results
According to Chen and Levi-Montalcini (1969, 1970), the brains or ganglia showed axonal outgrowth by the end of the 2nd day of culture. The
outgrowing axons branched repeatedly, and formed connections with
each other and with the outgrowth of other explants. A small enlargeFig. 3. Photomicrograph (Nomarski) of living 3-month-old culture of dissociated
nerve cells, thoracic ganglia (th), and foregut (j) explants from 16-day cockroach
embryos, Periptaneta americana. Nerve fibers branch out in large number from the
ganglion and interconnect with fibers produced by dissociated nerve cells. x 20.
From Levi-Montalcini and Aloe, "Neuronal nets and nerve cell interactions in vitro
in insect systems:' In Vitro 8, pp184. Copyright © 1972 by the Society for In Vitro
Biology (formerly the Tissue Culture Association). Reproduced with permission
of the copyright owner