American Society of Hematology

Figure 1

$Figure 1. PACSIN2 associates with FlnA in platelets. (A) Human and mouse platelet lysates were subjected to SDS-PAGE and probed for PACSIN2. (B) Increasing volumes of WT and Flna-null mouse platelet lysates were compared with known amounts of GST-PACSIN2 and probed for PACSIN2. The graph plots the number of PACSIN2 moles against platelet numbers obtained from 2 immunoblots. The amount of PACSIN2 per platelet was determined as the ratio between the first-order equation slopes best fitting the data. (C) Human platelets were activated or not with 10 µg/mL CRP for 2 minutes at 37°C and lysed, and PACSIN2 (P2) and FlnA were immunoprecipitated. A control rabbit IgG was used as negative control for specificity. Immunoprecipitates were subjected to SDS-PAGE and probed for FlnA. (D) Mouse platelets were activated with 1 U/mL thrombin (T) for 2 minutes at 37°C and lysed in PHEM buffer. Triton X-100 HSS and HSP fractions were collected by centrifugation of platelet lysates at 100 000g, subjected to SDS-PAGE, and probed for PACSIN2 and FlnA. CL, cell lysate.$

PACSIN2 associates with FlnA in platelets. (A) Human and mouse platelet lysates were subjected to SDS-PAGE and probed for PACSIN2. (B) Increasing volumes of WT and Flna-null mouse platelet lysates were compared with known amounts of GST-PACSIN2 and probed for PACSIN2. The graph plots the number of PACSIN2 moles against platelet numbers obtained from 2 immunoblots. The amount of PACSIN2 per platelet was determined as the ratio between the first-order equation slopes best fitting the data. (C) Human platelets were activated or not with 10 µg/mL CRP for 2 minutes at 37°C and lysed, and PACSIN2 (P2) and FlnA were immunoprecipitated. A control rabbit IgG was used as negative control for specificity. Immunoprecipitates were subjected to SDS-PAGE and probed for FlnA. (D) Mouse platelets were activated with 1 U/mL thrombin (T) for 2 minutes at 37°C and lysed in PHEM buffer. Triton X-100 HSS and HSP fractions were collected by centrifugation of platelet lysates at 100 000g, subjected to SDS-PAGE, and probed for PACSIN2 and FlnA. CL, cell lysate.

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