Analysis of AA/SAA reactivity in peripheral blood monocytes by confocal microscopy showed immunoreactivity in 5% of the monocytes isolated from a mouse with AA-amyloidosis (A).

<p>There was no reactivity present in monocytes recovered from a mouse given one AgNO₃ injection 48 hrs prior to isolation (B) or in monocytes isolated from untreated mice (C). The used rabbit antiserum recognizes both protein AA and SAA and was visualized by goat anti rabbit Alexa488-cojugated IgG. Cell nuclei were labeled with TO-PRO3. Bar 10 um.</p

Analysis of AEF activity in peripheral blood monocytes isolated from mice with AA-amyloid induced by AEF.

<p>AA-amyloid was induced in nine mice (G1–G9) by an i.v. injection of 0.1 ml AEF with concomitant s.c. injection of 0.2 ml 1% silver nitrate day 1, 7, 14, 21 and 28 and the mice were sacrificed on day 35. The presence of amyloid in the spleen was verified by Congo red staining. Peripheral blood monocytes were isolated, sonicated and re-introduced into the blood circulation of new groups of healthy mice (H1–H8). These mice received inflammatory stimuli day 1, 7 and 14 and were sacrificed day 16. The presence of amyloid was analysed in spleen sections after Congo red staining. Mice in group H9 received sonicated monocytes without subsequent inflammatory stimuli and group H10 received monocytes isolated from untreated mice and subsequent inflammatory stimuli on day 1, 7 and 14 and were sacrificed day 16 (group H10).</p

Spleen amyloid deposits stained with Congo red.

<p>(A) The amyloid appears pink and is localized to the perifollicular zone. (B) The identical area exhibits green birefringence in polarized light. Amyloid is indicated by arrows.</p

Analysis of AEF activity in peripheral blood monocytes isolated from mice with AA amyloidosis.

<p>The table presents detailed information on animals in group H 1–8. AA-amyloidosis was induced by i.v. injection of AEF and 0.2 ml 1% silver nitrate injections on day 1, 7, 14, 21 and 28. The animals were sacrificed on day 35. Blood was collected and amyloid was verified in spleen sections after Congo red staining. Isolated peripheral blood monocytes were injected into new animals, group H 1–8, and silver nitrate was given day 1, 7 and 14.The animals were sacrificed day 16 and the presence of amyloid was studied in spleen after Congo red staining.</p

SAA 1, SAA 2 and SAA 3 mRNA expression in peripheral blood monocytes were analysed with PCR.

<p>Cells were isolated from mice that developed AA-amyloid after AEF and AgNO₃ injections or from mice that received AEF or AgNO₃ injections only or from untreated mice. Expression of the amyloid-prone SAA 1 or non-amyloidogenic SAA 2 was absent in all monocyte preparations. SAA 3 mRNA was detected in all cells independent of treatment. Mouse liver cDNA was used as a positive control. The PCR products were separated on a 1.6% agarose gel.</p