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Method Article
A method for ameliorating autoimmune disease by passive transfer of IVIg-primed leukocytes
Andrew R Crow
commented on 20 July, 2006
In order to accurately reproduce our data, please note the following critical update regarding CD11c+ cell isolation: As of February 2006, StemCell Technologies have changed their kit (same name and same catalogue # but different components) and have added “mouse FcR blocker” to the “PE selection cocktail” (previously it was not part of the PE selection cocktail but was supplied as a separate reagent, which we did not add in our experiments), thus as of today, it is no longer an option to leave this FcR blocker antibody (anti-CD16/32) out of the assay. From this point forward, researchers purchasing this kit will not be able to reproduce our results. The company has indicated that they may manufacture a custom kit lacking the FcR blocker. In the interim, we advise researchers to use a CD11c selection kit which does not contain any antibodies which react with an FcR. Formally, please note the following clarification in the Cell Preparation step of the Procedure section: Step 2A (i) DO NOT add Mouse FcR blocker (antibody against mouse CD16/32) supplied with some versions of the dendritic cell isolation kit. Although the addition of FcR-blocker is recommended in the manufacturer’s instructions, this reagent will engage both activating and inhibitory Fc[gamma]Rs on the DC cells during purification leading to DCs which will subsequently be refractory to stimulation via IVIg as well as other Fc[gamma]R-specific crosslinking regimes. In cases where the DCs are from mice that are Fc[gamma]RIIB /, the addition of FcR blocker may cause DC activation rather than inducing a refractory state. Thus, If FcR blocker is added to the DCs during the purification process, subsequent dendritic cell priming will not work as expected.
Andrew R Crow
commented on 20 July, 2006
In order to accurately reproduce our data, please note the following critical update regarding CD11c+ cell isolation: As of February 2006, StemCell Technologies have changed their kit (same name and same catalogue # but different components) and have added “mouse FcR blocker” to the “PE selection cocktail” (previously it was not part of the PE selection cocktail but was supplied as a separate reagent, which we did not add in our experiments), thus as of today, it is no longer an option to leave this FcR blocker antibody (anti-CD16/32) out of the assay. From this point forward, researchers purchasing this kit will not be able to reproduce our results. The company has indicated that they may manufacture a custom kit lacking the FcR blocker. In the interim, we advise researchers to use a CD11c selection kit which does not contain any antibodies which react with an FcR. Formally, please note the following clarification in the Cell Preparation step of the Procedure section: Step 2A (i) DO NOT add Mouse FcR blocker (antibody against mouse CD16/32) supplied with some versions of the dendritic cell isolation kit. Although the addition of FcR-blocker is recommended in the manufacturer’s instructions, this reagent will engage both activating and inhibitory Fc[gamma]Rs on the DC cells during purification leading to DCs which will subsequently be refractory to stimulation via IVIg as well as other Fc[gamma]R-specific crosslinking regimes. In cases where the DCs are from mice that are Fc[gamma]RIIB /, the addition of FcR blocker may cause DC activation rather than inducing a refractory state. Thus, If FcR blocker is added to the DCs during the purification process, subsequent dendritic cell priming will not work as expected.
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CURRENT STATUS: Posted
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Posted 30 Jun, 2006
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Associated Publications
Intravenous immunoglobulin ameliorates ITPgamma via activating Fc receptors on dendritic cells
by Siragam, V. et al.
Nature Medicine (23 May, 2006)
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This is a preprint. It has not completed peer review.
Method Article
A method for ameliorating autoimmune disease by passive transfer of IVIg-primed leukocytes
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 30 Jun, 2006
Community comments: 2
Metrics
Comments: 2
HTML Views: ...
Andrew R Crow
commented on 20 July, 2006
In order to accurately reproduce our data, please note the following critical update regarding CD11c+ cell isolation: As of February 2006, StemCell Technologies have changed their kit (same name and same catalogue # but different components) and have added “mouse FcR blocker” to the “PE selection cocktail” (previously it was not part of the PE selection cocktail but was supplied as a separate reagent, which we did not add in our experiments), thus as of today, it is no longer an option to leave this FcR blocker antibody (anti-CD16/32) out of the assay. From this point forward, researchers purchasing this kit will not be able to reproduce our results. The company has indicated that they may manufacture a custom kit lacking the FcR blocker. In the interim, we advise researchers to use a CD11c selection kit which does not contain any antibodies which react with an FcR. Formally, please note the following clarification in the Cell Preparation step of the Procedure section: Step 2A (i) DO NOT add Mouse FcR blocker (antibody against mouse CD16/32) supplied with some versions of the dendritic cell isolation kit. Although the addition of FcR-blocker is recommended in the manufacturer’s instructions, this reagent will engage both activating and inhibitory Fc[gamma]Rs on the DC cells during purification leading to DCs which will subsequently be refractory to stimulation via IVIg as well as other Fc[gamma]R-specific crosslinking regimes. In cases where the DCs are from mice that are Fc[gamma]RIIB /, the addition of FcR blocker may cause DC activation rather than inducing a refractory state. Thus, If FcR blocker is added to the DCs during the purification process, subsequent dendritic cell priming will not work as expected.
Andrew R Crow
commented on 20 July, 2006
In order to accurately reproduce our data, please note the following critical update regarding CD11c+ cell isolation: As of February 2006, StemCell Technologies have changed their kit (same name and same catalogue # but different components) and have added “mouse FcR blocker” to the “PE selection cocktail” (previously it was not part of the PE selection cocktail but was supplied as a separate reagent, which we did not add in our experiments), thus as of today, it is no longer an option to leave this FcR blocker antibody (anti-CD16/32) out of the assay. From this point forward, researchers purchasing this kit will not be able to reproduce our results. The company has indicated that they may manufacture a custom kit lacking the FcR blocker. In the interim, we advise researchers to use a CD11c selection kit which does not contain any antibodies which react with an FcR. Formally, please note the following clarification in the Cell Preparation step of the Procedure section: Step 2A (i) DO NOT add Mouse FcR blocker (antibody against mouse CD16/32) supplied with some versions of the dendritic cell isolation kit. Although the addition of FcR-blocker is recommended in the manufacturer’s instructions, this reagent will engage both activating and inhibitory Fc[gamma]Rs on the DC cells during purification leading to DCs which will subsequently be refractory to stimulation via IVIg as well as other Fc[gamma]R-specific crosslinking regimes. In cases where the DCs are from mice that are Fc[gamma]RIIB /, the addition of FcR blocker may cause DC activation rather than inducing a refractory state. Thus, If FcR blocker is added to the DCs during the purification process, subsequent dendritic cell priming will not work as expected.
Associated Publications
Intravenous immunoglobulin ameliorates ITPgamma via activating Fc receptors on dendritic cells
by Siragam, V. et al.
Nature Medicine (23 May, 2006)