Difference between revisions of "Calabria 2017 MiP2017 WG4"
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{{Abstract | {{Abstract | ||
|title=[[File:CalabriaE.jpg|left|90px|Calabria Elisa]] | |title=[[File:CalabriaE.jpg|left|90px|Calabria Elisa]] PBMCS purification and mitochondrial function assay: monitoring preparative steps and comparison of protocols. | ||
PBMCS purification and mitochondrial function assay: monitoring preparative steps and | |||
comparison of protocols. | |||
|info=[[MiP2017]] | |info=[[MiP2017]] | ||
|authors=Calabria E, de Jong V, Tarperi C, Schena F | |authors=Calabria E, de Jong V, Tarperi C, Schena F | ||
Line 12: | Line 10: | ||
function. We have monitored all the steps of the PBMCs purification procedure with a Sysmex | function. We have monitored all the steps of the PBMCs purification procedure with a Sysmex | ||
XN-1000 hematology analyzer. This instruments allows a high degree of accuracy of platelets | XN-1000 hematology analyzer. This instruments allows a high degree of accuracy of platelets | ||
counts, also using a specific fluorescent reagent | counts, also using a specific fluorescent reagent [1]. The data collected show | ||
that we achieve 97% depletion of platelets after two washing steps in RPMI, and we collect the | that we achieve 97% depletion of platelets after two washing steps in RPMI, and we collect the | ||
30% of the initial amount of PBMCs. | 30% of the initial amount of PBMCs. | ||
One of the main aim of WG4 is also protocols in the harmonization to SUIT reference protocols | One of the main aim of [[WG4]] is also protocols in the harmonization to SUIT reference protocols | ||
(RP1 and RP2). Since we are not currently running exactly the suggested RP protocols, we tried to | (RP1 and RP2). Since we are not currently running exactly the suggested RP protocols, we tried to verify if at least some common functional steps of the protocol are comparable to RP1 and RP2 protocols. Preliminary data show that at least in some common steps there is no significant | ||
verify if at least some common functional steps of the protocol are comparable to RP1 and RP2 | |||
protocols. Preliminary data show that | |||
difference between the old protocol and the suggested RP1 and protocol in PBMCs human cells, | difference between the old protocol and the suggested RP1 and protocol in PBMCs human cells, | ||
further experiments are needed to evaluate RP2. | further experiments are needed to evaluate RP2. | ||
|editor=[[Beno M]], | |editor=[[Beno M]], [[Kandolf G]], | ||
|mipnetlab=IT Verona Calabria E | |mipnetlab=IT Verona Calabria E | ||
}} | }} | ||
{{Labeling}} | {{Labeling | ||
|tissues=Blood cells | |||
}} | |||
== References == | == References == | ||
:::: Wada A | ::::# Wada A, Takagi Y, Kono M, Morikawa T (2015) Accuracy of a new platelet count system (PLT-F)depends on the staining property of its reagents. PLOS ONE 10:1-13. |
Latest revision as of 14:00, 27 March 2018
PBMCS purification and mitochondrial function assay: monitoring preparative steps and comparison of protocols. |
Link: MiP2017
Calabria E, de Jong V, Tarperi C, Schena F (2017)
Event: MiP2017
In our laboratory we are mainly focused on PBMCs blood cells. One of the main issues of working with PBMCs is the quality of the final batch of cells. In particular the contamination of platelets in the final sample could affect the respiratory profile of the sample when evaluating mitochondrial function. We have monitored all the steps of the PBMCs purification procedure with a Sysmex XN-1000 hematology analyzer. This instruments allows a high degree of accuracy of platelets counts, also using a specific fluorescent reagent [1]. The data collected show that we achieve 97% depletion of platelets after two washing steps in RPMI, and we collect the 30% of the initial amount of PBMCs.
One of the main aim of WG4 is also protocols in the harmonization to SUIT reference protocols (RP1 and RP2). Since we are not currently running exactly the suggested RP protocols, we tried to verify if at least some common functional steps of the protocol are comparable to RP1 and RP2 protocols. Preliminary data show that at least in some common steps there is no significant difference between the old protocol and the suggested RP1 and protocol in PBMCs human cells, further experiments are needed to evaluate RP2.
β’ Bioblast editor: Beno M, Kandolf G
β’ O2k-Network Lab: IT Verona Calabria E
Labels:
Tissue;cell: Blood cells
References
- Wada A, Takagi Y, Kono M, Morikawa T (2015) Accuracy of a new platelet count system (PLT-F)depends on the staining property of its reagents. PLOS ONE 10:1-13.