Anti mouse ly6g clone 1a8

Two methods were used to deplete neutrophils in mice. For antibody-based depletion, WT mice were injected i.p. with anti-mouse Ly6G clone 1A8 (Bio X Cell) supernatant containing 0.5 mg protein every 48 hr for 1 month. Control animals were injected with isotype-matched normal control antibody. For genetic neutrophil ablation, diphtheria toxin receptor (DTR)-expressing Rosa26iDTR mice (Buch et al., 2005 (link); Saito et al., 2001 (link)) (The Jackson Laboratory) were bred onto the Mrp8-Cre genetic background (The Jackson Laboratory). Cre expression in MRP-expressing cells, mainly granulocytes, will activate DTR expression in only MRP-expressing cells. At the age of 3 months, the Rosa26iDTR/Mrp8-Cre mice and WT control littermates (Rosa26iDTR) were treated with diphtheria toxin (DT, 20 ng/g BW, i.p., daily for 3 days) to induce Mrp8⁺ cell depletion.
Clodronate-liposome (YEASEN, 40337ES05) was used to deplete mouse macrophage. WT mice were injected i.v. with 100 μL of Clodronate-liposome per 10 g of body weight. 24 hr later, mice were injected i.v. with 100 μL of Clodronate-liposome per 10 g of body weight again. The operation was repeated every 72 hr after the second injection for 1 month. Neutrophil and macrophage depletion were confirmed by neutrophil and macrophage count in all experimental animals.

This study is in compliance with the ARRIVE guidelines for in-vivo studies carried out on animals. Pregnant MRL/lpr and C57BL/6 J mice, aged 8–10 weeks, were purchased from Shanghai Model Organisms Center, Inc. The case and wild-type (WT) mice were randomly divided into a test group and a control group on the day after the vaginal plug was observed. The mice in the test group received an intraperitoneal injection of 200 µg anti-mouse Ly6G (clone 1A8; BioXcell, West Lebanon, USA) once every 4 days, for a total of 4 injections. The control group was injected with an equivalent volume of 0.9% normal saline (NS) following the same protocol as that of the anti-mouse Ly6G injection. There were 4 mice in WT-Ly6G group, 6 mice in MRL/lpr-Ly6G group, 5 mice in WT-NS group and 6 mice in MRL/lpr-NS group. Mice were weighed every day. On days 16–18, their placentas were collected, and the weights of the fetal mice were measured. To accurately compare the birth weights of the different groups, the fetal body weights were measured using the following equation: $$\text{The fetal body weight ratio}=\frac{\text{fetal birth weight}}{\text{maternal weight}\left(\text{on the day vaginal plug was observed}\right)}$$
Mice that died or had early pregnancy failure were excluded from the analysis.