Matt Gethers/20.380 HIV Project/Technical Paper/In Vitro Tests.doc: Difference between revisions

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In Vitro Tests
==In Vitro Tests==
-Genomic Testing of Stem cells
*Genomic Testing of Stem cells
oPCR, RT-PCR to see if things get added and expressed
**PCR, RT-PCR to see if things get added and expressed
-Proteomic Testing of Stem cells
*Proteomic Testing of Stem cells
oYes siRNA
**Yes siRNA
oNo RNAses, CD4+
**No RNAses, CD4+
-Genomic Testing of RBCs
*Genomic Testing of RBCs
oAka we want to see nothing in an RT-PCR or PCR
**Aka we want to see nothing in an RT-PCR or PCR
-Proteomic Testing of RBCs
*Proteomic Testing of RBCs
oLook for CD4+ (how much?), proteases
**Look for CD4+ (how much?), proteases
-Differentiation
*Differentiation
oMake sure the Stem cells only turn into RBCs
**Make sure the Stem cells only turn into RBCs
•Test by adding media that should trigger other differentiation
*Test by adding media that should trigger other differentiation
-Death Switch
*Death Switch
oMake sure our ‘death switch’ will kill only our RBCs/HSCs, not normal RBCs/HSCs  
**Make sure our ‘death switch’ will kill only our RBCs/HSCs, not normal RBCs/HSCs  
oDetermine concentration necessary to kill cells with death switch (whoot cytotoxicity)
**Determine concentration necessary to kill cells with death switch (whoot cytotoxicity)
-Functioning of RBCs
*Functioning of RBCs
oDeformation tests – indicative of clearing from blood
**Deformation tests – indicative of clearing from blood
oStill able to carry oxygen?
**Still able to carry oxygen?
-HIV Infection controlled
*HIV Infection controlled
oSee if the HSCs get infected (two fold: 1 see if they get infected, 2 see if the siRNAs work)
**See if the HSCs get infected (two fold: 1 see if they get infected, 2 see if the siRNAs work)
oSee if the RBCs get infected
**See if the RBCs get infected
•How many virions per cell
*How many virions per cell
•Under normal conditions
*Under normal conditions
•If at max capacity of virions, do cells explode
*If at max capacity of virions, do cells explode
oIs the virion released able to infect?
**Is the virion released able to infect?
•Do the virions get destroyed
*Do the virions get destroyed
•Is it by siRNA or RNAses primarily?
*Is it by siRNA or RNAses primarily?
-Co-culture
*Co-culture
oPut engineered RBCs and T cells together
**Put engineered RBCs and T cells together
•Who gets infected?
*Who gets infected?
•How many RBCs are necessary per T cell
*How many RBCs are necessary per T cell
•How many eRBCs versus normal RBCs?
*How many eRBCs versus normal RBCs?
•If RBCs filled with virus (but no free floating virus) are added to T cell culture, do T cells get infected?
*If RBCs filled with virus (but no free floating virus) are added to T cell culture, do T cells get infected?
-HIV effects
===HIV effects===
oDo we see resistance in HIV?
*Do we see resistance in HIV?
oLook for evolution of strain<nowiki><nowiki>Insert non-formatted text here</nowiki><nowiki>Insert non-formatted text here</nowiki></nowiki>
**Look for evolution of strain

Latest revision as of 19:30, 15 April 2009

In Vitro Tests

  • Genomic Testing of Stem cells
    • PCR, RT-PCR to see if things get added and expressed
  • Proteomic Testing of Stem cells
    • Yes siRNA
    • No RNAses, CD4+
  • Genomic Testing of RBCs
    • Aka we want to see nothing in an RT-PCR or PCR
  • Proteomic Testing of RBCs
    • Look for CD4+ (how much?), proteases
  • Differentiation
    • Make sure the Stem cells only turn into RBCs
  • Test by adding media that should trigger other differentiation
  • Death Switch
    • Make sure our ‘death switch’ will kill only our RBCs/HSCs, not normal RBCs/HSCs
    • Determine concentration necessary to kill cells with death switch (whoot cytotoxicity)
  • Functioning of RBCs
    • Deformation tests – indicative of clearing from blood
    • Still able to carry oxygen?
  • HIV Infection controlled
    • See if the HSCs get infected (two fold: 1 see if they get infected, 2 see if the siRNAs work)
    • See if the RBCs get infected
  • How many virions per cell
  • Under normal conditions
  • If at max capacity of virions, do cells explode
    • Is the virion released able to infect?
  • Do the virions get destroyed
  • Is it by siRNA or RNAses primarily?
  • Co-culture
    • Put engineered RBCs and T cells together
  • Who gets infected?
  • How many RBCs are necessary per T cell
  • How many eRBCs versus normal RBCs?
  • If RBCs filled with virus (but no free floating virus) are added to T cell culture, do T cells get infected?

HIV effects

  • Do we see resistance in HIV?
    • Look for evolution of strain
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