1. Isaacs and Lindenmann discovered the presence of interferons by incubating he
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1. Isaacs and Lindenmann discovered the presence of interferons by incubating heat-killed influenza virus (MEL strain) with the chorioallantoic membranes of embryonated eggs. We went through some of their primary data (table below) in class. To review, they incubated with heated MEL for the length of time indicated in the "primary incubation" column. After washing, they continued to incubate for additional time (secondary incubation") followed by direct challenge with live MEL. They then measured how much virus was able to grow out. TABLE 1. EFFECT OF VARYING TIMES AND TEMPERATURES OF INCUBATION ON THE INTERFERING ACTIVITY OF HEATED MEL geometric mean HA titre (log2) 3.9 %of control titre 15 interfering virus primary incubation secondary incubation 60 a.d. of heated MEL 15 in at 37° C 24 h at 37°C 60 a.d. of heated MEL 15 min at 37 C 24 h at 2°C 60 a.d. of hcated MEL1h at 37°C 60 a.d. of heated MEL h at 37°C 60 a.d. of heated MEL4 h at 37° C 60 a.d. of heated MEL 4h at 37 C 60 a.d. of heated MEL 24h at 37°O buffer control buffer control >100 23 h at 37°C 23 h at 2°C 20 h at 37°C 20 h at 2 C 4-0 5-8 2-3 3.6 94 20 24 h at 37°C 24 h at 2°C nil nil ni 5-6 5.9 100 100 (a) Based upon concepts discussed in class, how can a killed, inactivated virus induce the production of interfering activity (termed "interferons")? [2 pts.] (b) We know that interferons are secreted factors that act on neighboring cells. Devise an experiment using the experimental system of Isaacs and Lindenmann to show that interferons are indeed secreted proteins. [3 pts.] (c) Look at the data generated for the 15 min and 4 hour primary incubations followed by secondary incubation at 2°C (lines 2 and 6 of data). The former had no negative impact on virus replication whereas the latter greatly decreased viral titers. Provide an explanation for these findings. [3 pts.]Explanation / Answer
Hi,
The interferon are the proteins or signaling molecules. These are produced by the host cell in response to viral infection. The viral when attacks the host , the host detects it using its specific antigenic particles, such as glycoproteins, viral mRNA…etc. These are then processed by the Toll like receptors on the surface of the host cell and series events follow to produce interferon. In order to detect the viral infection, a cell needs only the viral particles, and hence the heat inactivated viral can also induce interferon.
If the interferon is indeed protein, then we can devise a simple experiment to test this. After the washed membrane is treated with live virus, the cell suspension can be tested using specific antibody probes for interferons. If the binding is successful, it also means that the interferons are proteins. We can also do SDS_PAGE of the interferon suspensions
The lane 2 and lane 4 differ in the incubation time and show difference in interference. This probably indicates that the metabolic process in the membrane requires at least 4h of incubation at 37C. Interference can be established only after this 4h of incubation. Even though 15 min of primary incubation is sufficient provided with 24h secondary incubation, the 4h incubation greatly enhances the effect.
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