The use of radioactive phosphorus in studies of chick embryo infections with vaccinia and influenza virus

121 Pages·4.493 MB·English

by  Altenburg, Luolin Storey

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THE RICE INSTITUTE
the use of radioactive phosphorus in studies of chick embryo
INFECTIONS WITH VACCINIA AND INFLUENZA VIRUS
Luolin Storey Altenburg
A THESIS
SUBMITTED TO THE FACULTY
IN PARTIAL FULFILLMENT OF THE
REQUIREMENTS FOR THE DEGREE OF
DOCTOR OF PHILOSOPHY
Houston, Texas
May, 1951
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Acknowledgments
This investigation has been completed under the
direction of Professor Edgar Altenburg, to whom the author
wishes to express sincere appreciation for valuable assistance
in outlining the problem and designing the experiments. Acknowl­
edgment Is also made of the cooperation of Professor Roy 7.
Talmage, director of the radiobiology laboratory of The Rice
In stitu te. Thanks are due to Professor A. C. Chandler for
helpful suggestions cna also to Professor H. D. Brunk for
assistance in regard to the sta tistic a l treatm ent of the data
herein presented. The author is likew ise indebted to the
following students for capable and careful assistance in the
actual performance of the experiments: Robert N. Baker,
John E. Simmons, Sophy Silversteen, and David Aldrich.
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TABLE OF CONTENTS
INTRODUCTION.....................................................................................................................................................
REVIEW OF LITERATURE................................................................................................................................4
STUDIES ON VACCINIA VIRUS...................................................................................................................10
A. M aterials and Methods........................................... 10
1. Inoculation of Embryos .......................... 10
2. Harvesting of Fluids and Membranes..................... 12
3. D ialysis..................... 14
4. Techniques Involving the Use of Radioactive Phosphorus.........................16
B. Experiments in Which P^ 2 WasF ed to Laying H e n s .....................................1 8
C. Experiments in Which p32 Was Injected into the Yolk.......................................20
D. Experiments in Which P^ 2 Was Injected into the Albumen................................22
1. p32 in Normal allantoic Fluid................................. 24
2. P®2 in A llantoic Fluids after 72 Hours of Virus Growth......................25
3. P ^ in A llantoic Fluids after from 1 to 30 Hours of Virus
Growth.........................................................................................................................................27
4. P ^ in Chorio-allantoic M em branes...................................................................30
E. Discussion............................ 43
F. Summary............................................... 46
STUDIES ON INFLUENZA VIRUS................................................................................................................48
A. M aterials and Methods............................ ....4 9
1. Preservation of the V irus................ 49
2. Inoculation of Embryos by the A llantoic Route............................................49
3. Inoculation of Embryos by the Amniotic R o u te .................................5 1
4. Harvesting of Fluids, Membranes, and E m b ry o s............................5 2
5. The Hemagglutination Technique.................................................................................55
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B. Experiments in Which Influenza Virus Was Inoculated by the
A llantoic Route................................................................................................... .57
32
1 . P in C horio-allantoic Membranes.............................................. .57
2. P^2 in A llantoic Fluids after 24 Hours of Virus Growth .69
3. P ^ in A llantoic Fluids after 40 Hours of Virus Growth .77
C. Experiments in Which Influenza Virus Was Inoculated by the
Amniotic Route..................................................................................................... .83
D. Discussion............................................................................................................... .89
E. Summary ............. .98
3IBLI0G.lf PHY............................................................................................................................. 103
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The Use of Radioactive Phosphorus in Studies of Chiok Embryo Infections
w ith Vaccinia and Influenza Viruses
INTRODUCTION
The need for a simple and speedy method of demonstrating the
presence of virus in laboratory cultures has long been apparent, both,
to the medical profession and to the research worker* A few viruses
oause v isib le lesions or characteristic aggregates which are large
enough to be seen as oell inclusion bodies in infected m aterial, but
many viruses must be detected by other means, most of which have been
borrowed from bacteriology. I t should be mentioned here th at bio­
chemical methods (involving pH changes and turbidity determ inations,
for example) have failed up to the present time to d ifferentiate be­
tween v iral and noxrviral m aterial, except in one instance reported
by A tlas and H ottle (1948) in the case of a recently isolated oomnon
cold virus* Exoept for animal infection (in whioh disease symptoms
or death are looked for in susceptible hosts and the agent recovered
from thorn), most other tests are serological in nature; th at is, they
are based on the fact th at seme viruses oause the production of demon­
strable antibodies. There are three serologioal tests now in use for
human diseases* (1 ) neutralization, (2 ) complement fixation, and
(3) hemagglutination-inhibition tests* The neutralization te st in­
volves the injection into susceptible host animals of serum suspected
of containing antibodies against a particular virus along with a su it­
able dilution of the virus itself* If the animals are protected or
p artially protected against the virus, a ten tative identification is
established. However, the serum being tested may simply have contained
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