Antibodies play a critical role in adaptive immune responses and serve as key components in disease diagnosis and treatment. These molecules exhibit dynamic post-translational modifications (PTMs), such as glycosylation and phosphorylation, which regulate their effector functions. To date, nearly all of our knowledge about antibody repertoires has come from B cell receptor (BCR) sequencing (BCR-seq), which facilitates the profiling of clonal composition and the tracing of maturation trajectories within B-cell repertoires. However, circulating antibodies found in bodily fluids—such as serum, saliva, milk, mucosal secretions, and cerebrospinal fluid—exhibit diversities and specificities beyond what BCR-seq alone can predict. Therefore, identifying and quantifying antibody clonotypes at the protein level could enhance diagnosis, prognosis, and treatment strategies in personalized medicine. The critical gap between genotype and phenotype necessitates complementary methodologies that enable the direct characterization of antibody proteins in their native functional states. Mass spectrometry (MS)-based antibody repertoire sequencing (Ab-seq) is currently the only feasible approach for this task and primarily includes database-dependent methods—such as bottom-up, middle-down, and top-down approaches—as well as database-independent de novo sequencing technology. These strategies enable multi-level, high-precision characterization ranging from peptides and domains to intact antibody molecules. Unlike the shotgun strategy commonly used in routine proteomics, obtaining full sequences of all antibodies presents unique challenges. It requires specialized methodological adaptations to address issues related to dynamic range, sequence variation, and sample complexity. This review introduces the technical principles, methodological workflows, and recent applications of various mass spectrometry-based antibody repertoire sequencing (Ab-seq) strategies, with a focus on approaches designed to improve sequence coverage and identification accuracy. These include multi-enzyme digestion, hybrid fragmentation methods, and artificial intelligence-assisted de novo sequencing. By systematically comparing database-dependent techniques—such as bottom-up, middle-down, and top-down approaches—with database-independent de novo sequencing, this review outlines their respective advantages and limitations in terms of sample throughput, sequence coverage, post-translational modification characterization, and data analysis complexity. In addition, this review discusses emerging technological trends, including the integration of ion mobility separation, native mass spectrometry, and artificial intelligence-driven data interpretation, which are expected to enhance the depth and accuracy of antibody characterization. Although current methods continue to face challenges related to sample complexity, dynamic range, and unambiguous sequence variant assignment, we emphasize the importance of integrating BCR-seq and Ab-seq data to construct gene-protein association maps. These maps help validate sequence accuracy and facilitate epitope discovery. This dual-platform strategy helps bridge the gap between genotype and phenotype, thereby enhancing both the resolution and scope of antibody repertoire studies. Such an integrative approach also offers a valuable tool for therapeutic antibody development, structure-function analysis, and precise evaluation of vaccine efficacy.

Objective:To analyze the genetic characteristics of the VWF gene c.7332G＞A nonsense mutation and explore its molecular pathogenesis.Methods:Phenotypic diagnosis of the proband was performed using VWF:Ag,VWF:RCo,FⅧ:C and multimeric analysis.The probands were genotyped by NGS whole-exome sequencing,and the sequencing results were validated by sanger sequencing.The family members were genotyped by Sanger sequencing.The VWF gene c.7332G＞A nonsense mutant plasmid was constructed.After transfection,the function of VWF gene c.7332G＞A mutant plasmid was verified at cell level in vitro.The mRNA level was detected by qRT-PCR,and the expression level of protein was detected by Western blot,the function of multimerization was verified by the multimeric analysis.Results:VWF:Ag and VWF:RCo were all less than 3％in the proband,and the multimeric analysis showed multimer deficiency.The proband was diagnosed as type 3 VWD.The homozygous nonsense mutation of VWF gene c.7332G＞A was detected by gene sequencing.The VWF mRNA level of the mutant plasmid was decreased,and the VWF protein expression in the cell supernatant was decreased,the mutant protein was truncated and the function of VWF multimerization was impaired.Conclusion:A homozygous mutation in exon 43 of VWF gene,c.7332G＞A,was responsible for the probands type 3 VWD in the proband.The mutation caused a decrease in the relative level of VWF mRNA and protein,and impaired the function of VWF multimerization.

Objective:To perform RHD gene detection on a blood sample with serological weak D phenotype.Methods:A specimen received by the People's Hospital of Zhijin County was serologically identified by the microcolumn gel method and saline method.RHD gene detection was conducted by the PCR-SSP method,and the full sequence determination of the 10 exons amplified was performed.The sequencing results were compared with the ISBT database to determine the genotype.Bioinformatics tool was used to predict the functional damage of mutant proteins,and Alphafold-3 was used for tertiary structural modeling of wild-type and mutant RhD proteins,and the structures of the two proteins were compared and analyzed to explore the reasons why mutations lead to weak serological manifestations.Results:The patient's genotype was identified as RHD*DV.5/RHD*01N.01 heterozygote,with the complete deletion of RHD genes on one chromosome,unable to express the D antigen.On the other chromosome,a G＞A mutation occurred at the 697th base of the 5th exon,resulting in a partial D phenotype.This mutation causes internal hydrogen bond changes at the 233 position of RhD protein,resulting in a change in the conformation of the protein,affecting binding to the corresponding antibody.Conclusion:The patient is a heterozygous mutant individual with RHD*DV.5/RHD*01N.01,exhibiting a partial D phenotype serologically.This variation is extremely rare and has been scarcely reported globally.

Objective:To establish a stable rat model of sequelae of pelvic inflammatory disease(SPID)with clinical characteristics,and to provide a reliable experimental model for the study of the pharmcological effect and mechanism of SPID.Methods:Twenty-four 7-week-old SD rats were divided into sham operation group,model-A(108 cfu/mL mixed bacterial solution,0.2 mL),model-B(109 cfu/mL mixed bacterial solution 0.2 mL),and model-C(108 cfu/mL E.coli 0.2 mL).The weight of the rat's uterine was weighed and the uterine index was calculated.The automatic hematology analyzer was used to detect the blood routine;hematoxylin-eosin staining(HE)and masson staining were used to detect uterine pathlogical changes in rats.Enzyme-linked immunosorbent assay(ELISA)was used to detect interleukin-1β(IL-1β),interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)in rat uterine tissue homogenates.Western blot was used to detect the expression of proteins related to NF-κB signaling pathway.Results:Compared with the sham operation group,the uterine index of model-A,model-B,and model-C were significantly increased(P＜0.05,P＜0.01).The levels of WBC and NE in the model-A increased significantly(P＜0.01).The level of LY in model-B decreased significantly(P＜0.01).The levels of IL-1β,TNF-α in model-A,model-B,and model-C were significantly increased(P＜0.01).The levels of IL-6 in model-A and model-B were significantly increased(P＜0.05,P＜0.01).The collagen volume fraction of model-A and model-B were significantly increased(P＜0.01).Mechanism study indicates that the expression levels of p-IKKβ/IKKβ,p-IκBα/IκBα and p-p65/p65 in model-A were significantly increased(P＜0.01),and the expression levels of IκBα/β-actin were significantly decreased(P＜0.01).The expression level of p-IKKβ/IKKβ in model-B was significantly increased(P＜0.01).Conclusions:A stable rat model of SPID that conforms to clinical characteristics can be successfully constructed by combining 0.2 mL of mixed bacterial solution with a concentration of 108 cfu/mL and mechanical injury.This modeling method intervened in the expression of the NF-κB inflammatory signaling pathway.

The ocean plays a critical role in the global carbon cycle,and base on the"dual carbon"goals,ocean carbon sinks have received widespread attention.Shellfish aquaculture is one of the most important sources of carbon sinks in fisheries,which has an important impact on the offshore carbon cy-cle.As global temperature rises and ocean acidification intensifies,the capacity of the ocean to absorb CO2 will change.However,the effects of high temperature on the physiology and transcriptome related to carbon metabolism in Mytilus coruscus are not clear enough.This study investigated the effects of high temperatures on the total carbon content,carbon metabolism,antioxidant-related enzyme activities,and the transcriptome of Mytilus coruscus.The results showed that high temperature significantly inhibited the activities of hexokinase and pyruvate kinase,increased carbonic anhydrase activity(P＜0.05),de-creased the ATP content of digestive glands(P＜0.05),and affected glycolysis and the tricarboxylic acid cycle,leading to a significant decrease in the mussel's ability to sequester carbon.High temperature re-sulted in significant(P＜0.05)increases in the levels of reactive oxygen species and malondialdehyde,and enhanced the activities of superoxide dismutase and catalase.Observations by transmission electron microscopy showed that high temperatures damaged the subcellular structure of the digestive gland in Mytilus coruscus,resulting in the shrinkage of the nucleolus,swelling of the endoplasmic reticulum,and a significant reduction in the mitochondrial cristae.Comparative transcriptomic analysis showed that the upregulated DEGs were mainly enriched in protein processing in the endoplasmic reticulum,antigen pro-cessing and presentation,and MAPK signaling pathway.The downregulated DEGs were mainly enriched in necroptosis,DNA replication,and the NF-kappa B signaling pathway.In antioxidant-related DEGs,the upregulated DEGs include vitamin K epoxide reductase,peroxidases,heat shock protein 105 kD,heat shock protein 70 kD,and superoxide dismutase;The downregulated DEGs mainly included NADPH oxidase,glutathione reductase,cytochrome b-245,cytochrome P450,and quinone reductase.The up-regulated genes enriched in the carbon metabolism pathway included chitinase,phosphatidylinositol 4,5-bisphosphate 3-kinase,phosphoenolpyruvate carboxykinase,galactokinase,and inositol trisphosphate 3-kinase.The downregulated genes included aldose-1-epimerase,carbonic anhydrase,galactose mutaro-tase,acyl-CoA synthetase,alcohol dehydrogenase,and hexokinase.In conclusion,high temperature has an inhibitory effect on the activities of enzymes and the expression of genes related to carbon metabolism in Mytilus coruscus.The results of this study are intended to provide a scientific basis for the healthy de-velopment of mussel aquaculture and the assessment of carbon sinks.

Objective:To establish a stable rat model of sequelae of pelvic inflammatory disease(SPID)with clinical characteristics,and to provide a reliable experimental model for the study of the pharmcological effect and mechanism of SPID.Methods:Twenty-four 7-week-old SD rats were divided into sham operation group,model-A(108 cfu/mL mixed bacterial solution,0.2 mL),model-B(109 cfu/mL mixed bacterial solution 0.2 mL),and model-C(108 cfu/mL E.coli 0.2 mL).The weight of the rat's uterine was weighed and the uterine index was calculated.The automatic hematology analyzer was used to detect the blood routine;hematoxylin-eosin staining(HE)and masson staining were used to detect uterine pathlogical changes in rats.Enzyme-linked immunosorbent assay(ELISA)was used to detect interleukin-1β(IL-1β),interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)in rat uterine tissue homogenates.Western blot was used to detect the expression of proteins related to NF-κB signaling pathway.Results:Compared with the sham operation group,the uterine index of model-A,model-B,and model-C were significantly increased(P＜0.05,P＜0.01).The levels of WBC and NE in the model-A increased significantly(P＜0.01).The level of LY in model-B decreased significantly(P＜0.01).The levels of IL-1β,TNF-α in model-A,model-B,and model-C were significantly increased(P＜0.01).The levels of IL-6 in model-A and model-B were significantly increased(P＜0.05,P＜0.01).The collagen volume fraction of model-A and model-B were significantly increased(P＜0.01).Mechanism study indicates that the expression levels of p-IKKβ/IKKβ,p-IκBα/IκBα and p-p65/p65 in model-A were significantly increased(P＜0.01),and the expression levels of IκBα/β-actin were significantly decreased(P＜0.01).The expression level of p-IKKβ/IKKβ in model-B was significantly increased(P＜0.01).Conclusions:A stable rat model of SPID that conforms to clinical characteristics can be successfully constructed by combining 0.2 mL of mixed bacterial solution with a concentration of 108 cfu/mL and mechanical injury.This modeling method intervened in the expression of the NF-κB inflammatory signaling pathway.

Ischemic stroke(IS)research has faced bottlenecks due to the limitations of conventional technologies in resolving cellular heterogeneity and spatiotemporal dynamics.The development of single-cell and spatial omics technologies provides revolutionary tools to break through these constraints.Single-cell omics technologies,by performing high-throughput sequencing on thousands of cells,reveal the high heterogeneity and dynamic state transitions of neurons,glial cells,immune cells,and others post-IS.For instance,microglia contain pro-inflammatory and anti-inflammatory functional subsets,while astrocytes exhibit distinct activation state spectra.Pseudotime analysis further reconstructs the fate trajectories of cells during the damage and repair processes.Spatial omics technologies,conversely,reconstruct spatial maps of gene expression through in situ capture,elucidating molecular gradients between the ischemic core,penumbra,and healthy brain regions,and enabling the analysis of critical cell-cell interaction net-works.Integrating the deep phenotyping capability of single-cell sequencing with the in situ localization information from spatial omics constitutes the current core strategy.This multimodal analysis allows for precise anchoring of cell subtypes to their spatial microenvironments,revealing their distribution patterns and functions,and constructing a more accurate atlas of cell-cell communication.This significantly ad-vances the refined dissection of IS mechanisms.This strategy has already accelerated the discovery of po-tential biomarkers and spatiotemporally specific therapeutic targets.Although challenges remain in sample preparation,data integration,and technical noise,future interdisciplinary collaboration,multi-omics in-tegration,and in-depth mining with artificial intelligence promise to comprehensively transform our under-standing of IS.Ultimately,it holds the potential to promote advances in its early diagnosis,precise sub-typing,and the development of individualized treatment strategies.

Aim To investigate the anti-acute lung injury(ALI)effect of Sterculiae Lychnophorae Semen(SLS)and its mechanism.Methods The main ac-tive components of SLS and their core targets and path-ways of action against ALI were obtained by network pharmacology methods.Subsequently,molecular doc-king technology and in vitro cellular experiments were applied for validation.Results A total of 19 core tar-gets were obtained,including HSP90AA1,CASP3,TNF,MAPK8 and MAPK14.The mechanisms may in-volve signaling pathways such as cancer,PI3K/Akt and MAPK.Molecular docking confirmed that the key targets of SLS formed a better binding activity with the relevant active ingredients.The in vitro results showed that SLS was able to protect the PM2.5-contaminated BEAS-2B cells,inhibit their NO,IL-1β and TNF-αlevels,and reduce the expression of p-p38 MAPK and p-JNK proteins.Conclusions The study successfully predicts the active ingredients,targets and signaling pathways of SLS against ALI,and in vitro experiments demonstrate that SLS might protect BEAS-2B cells from PM2.5 stimulus-induced inflammation and apoptosis by inhibiting the over-activation of p38 MAPK and JNK signaling pathways.

Objective:To develop a non-pharmacological nursing intervention program based on enhanced recovery after surgery (ERAS) and wrist-ankle acupuncture for postoperative nausea and vomiting (PONV) in gynecological laparoscopic surgery, and to evaluate its effectiveness.Methods:Through systematic literature retrieval, evidence screening, and expert consultation, a non-pharmacological intervention program for PONV based on ERAS and wrist-ankle acupuncture was established. Patients undergoing gynecological laparoscopic surgery in Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine from January to May 2025 were selected using convenience sampling. The observation group received the ERAS and wrist-ankle acupuncture-based intervention program, while the control group received sham acupuncture on the basis of routine nursing care. The Rhodes Index of Nausea, Vomiting, and Retching (R-INVR) , the Postoperative Patient Comfort Scale (PPCS) , adverse event incidence, and patient satisfaction were used to evaluate intervention effects.Results:A total of 96 patients completed the study, including 49 in the observation group and 47 in the control group. After the intervention, 13 patients in the observation group and 23 in the control group had R-INVR scores ≥4, with a statistically significant difference between groups ( P<0.05) . The PPCS score was (70.76±16.11) in the observation group and (61.09±12.24) in the control group, with a significant difference ( t=3.321, P<0.05) . Patient satisfaction was higher in the observation group than in the control group ( P<0.05) . During the treatment period, all patients had stable vital signs, and no serious adverse reactions or allergic responses to drugs or acupuncture were observed. Conclusions:The ERAS and wrist-ankle acupuncture-based non-pharmacological nursing intervention program is reasonable and effective. From the perspective of patient safety, it can reduce the severity of PONV in gynecological laparoscopic surgery and improve patient comfort and satisfaction.

Blood from a case of group B epidemic cerebrospinal meningitis identified in February 2024 in Ganzhou City,Jiangxi Province,and throat swabs from close contacts were collected for isolation and culture.The isolates were subjected to serogrouping,drug sensitivity testing,and whole genome sequencing and analysis,to provide a basis for epidemiological inves-tigation and clinical drug use.One strain of Neisseria meningitidis was isolated from the blood of the case and denoted group B.The MLST type was ST-7962,with no clonal group attribution.The phylogenetic tree showed that it was genetically close to the 1977 Shanghai carrier isolate(id-52231).Drug sensitivity results indicated that the strain was sensitive to 8 drugs:azithro-mycin,cefotaxime,minocycline,ceftriaxone,chloramphenicol,meropenem,rifampicin,and benzylpenicillin;resistant to cot-rimoxazole,levofloxacin,and ciprofloxacin;and showed an intermediate response to penicillin.This report describes the first case of ST-7962 group B meningoencephalitis found in Jiangxi Province.Monitoring of Neisseria meningitidis carriage,drug re-sistance,and molecular characteristics of strains in the healthy population in this region should be strengthened,to provide la-boratory support for the clinical use of medications,traceability,and control of the pathogen underlying meningoencephalitis infection.