Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: (A) Confocal images of endogenous eIF2B subunits localizing to cytoplasmic foci. U373 and SH-SY5Y cells were fixed in methanol, MO3.13 cells were fixed in 4%PFA and subjected to ICC with (left to right) anti-eIF2Bα, anti-eIF2Bβ, anti-eIF2Bγ, anti-eIF2Bδ, or anti-eIF2Bε primary antibodies and visualized using appropriate secondary antibodies conjugated to Alexa Fluor 594. (B) Co-localised eIF2Bα and eIF2Bγ in U373, MO3.13 and SH-SY5Y (i) – (iii). Size distribution of eIF2Bα foci co-localised and not co-localised with eIF2Bγ (n=3 counts of 30 cells with eIF2Bα localisation). (i) = U373 cells; (ii) = MO3.13 cells; (iii) = SH SY5Y cells (C) Percentage of cells that showed localisation, i.e., one or more foci of eIF2B subunits (n=3 counts of 100 cells) presented as mean ± SD. p Values derived from a two-way ANOVA test, followed by a Tukey’s multiple analysis, *p ≤ 0.05, ***p ≤ 0.001, ****p ≤ 0.0001. (D) Average number of small (<1µm 2 ) eIF2Bα, eIF2Bβ, eIF2Bγ, eIF2Bδ, or eIF2Bε foci per cell in U373, MO3.13 and SH-SY5Y cells, presented as mean ± SD (n=3 counts of 30 cells with localised foci). p Values derived from an ordinary one-way ANOVA test, followed by a Tukey’s multiple analysis, *p ≤ 0.05, **p ≤ 0.01. (E) Western Blot analysis of the levels of eIF2Bα expression in U373, MO3.13 and SH-SY5Y cells under untreated conditions. Levels of eIF2Bα were normalized to levels of β-actin. The average ratio of eIF2Bα protein to β-actin is shown under each cell line, standard deviation in brackets

Article Snippet: The following antibodies were used: eIF2α (Abcam ab5369; 1:500), phosho-eIF2α[ser51] (Abcam ab32157; 1:500), GAPDH (Cell Signalling #2118; 1:5,000), β-actin (Cell Signalling #3700; 1:2500), eIF2Bα (Proteintech 18010-1-AP; 1:500), eIF2Bβ (Proteintech 11034-1-AP; 1:500), eIF2Bγ (Santa Cruz sc-137248; 1:500), eIF2Bδ (Proteintech 11332-1-AP; 1:50), eIF2Bε(Sigma-Aldrich HPA064370; 1:500).

Techniques: Derivative Assay, Western Blot, Expressing, Standard Deviation

Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: (A) Confocal images of endogenous small and large cytoplasmic foci of eIF2Bα. U373 and SH-SY5Y cells were fixed in methanol, MO3.13 cells were fixed in 4%PFA and subjected to ICC with anti-eIF2Bα primary antibodies and visualized using appropriate secondary antibodies conjugated to Alexa Fluor 594. (B) Confocal images of endogenous eIF2Bα and eIF2Bγ in U373 cells fixed in methanol and subjected to ICC with anti-eIF2Bα and anti-eIF2Bγ primary antibodies visualized using appropriate secondary antibodies conjugated to Alexa Fluor 594 & 488. (C) Average number of large (>1µm 2 ) eIF2Bα, eIF2Bβ, eIF2Bγ, eIF2Bδ, or eIF2Bε foci per cell in U373, MO3.13 and SH-SY5Y cells, presented as mean ± SD (n=3 counts of 30 cells with localised foci). p Values derived from an ordinary one-way ANOVA test, followed by a Tukey’s multiple analysis, * p ≤ 0.05. (D) Western Blot analysis of the levels of eIF2Bα-ε expression in U373, MO3.13 and SH-SY5Y cells under untreated conditions. Levels of eIF2Bα-ε were normalized to levels of β-actin. The average ratio of eIF2Bα protein to β-actin is shown under each cell line, standard deviation in brackets

Article Snippet: The following antibodies were used: eIF2α (Abcam ab5369; 1:500), phosho-eIF2α[ser51] (Abcam ab32157; 1:500), GAPDH (Cell Signalling #2118; 1:5,000), β-actin (Cell Signalling #3700; 1:2500), eIF2Bα (Proteintech 18010-1-AP; 1:500), eIF2Bβ (Proteintech 11034-1-AP; 1:500), eIF2Bγ (Santa Cruz sc-137248; 1:500), eIF2Bδ (Proteintech 11332-1-AP; 1:50), eIF2Bε(Sigma-Aldrich HPA064370; 1:500).

Techniques: Derivative Assay, Western Blot, Expressing, Standard Deviation

Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: (A) Average number of eIF2Bα foci per cell in a population of 30 cells with localised eIF2Bα, untreated, DMSO and Tg 1μM 1h treatment (N=3). Data was analysed using one-way ANOVA followed by post-hoc Tukey’s test for multiple comparisons. Error bars: ± s.d. (N=3). *p≤0.05; **p≤0.01. (i) small foci <1µm 2 ; (ii) = large foci ≥1µm 2 (B) Average number of eIF2Bε foci per cell in a population of 30 cells with localised eIF2Bε, untreated, DMSO and Tg 1μM 1h treatment (N=3). Data was analysed using one-way ANOVA followed by post-hoc Tukey’s test for multiple comparisons. Error bars: ± s.d. (N=3). *p≤0.05; **p≤0.01. (i) small foci <1µm 2 ; (ii) = large foci ≥1µm (C) (i)Western Blot analysis of the levels of eIF2Bα, eIF2Bε p-eIF2α and total eIF2α expression in U373, MO3.13 and SH-SY5Y cells, following DMSO and Tg 1μM 1h treatment. Levels of (ii) eIF2Bα and (iii) eIF2Bε were normalized to levels of β-actin (N=3). (iv) Levels of p-eIF2α were normalized to levels of total eIF2α (N=3). Data was analysed using two-way ANOVA followed by post-hoc Tukey’s test for multiple comparisons. Error bars: ± s.d. (N=3). * p ≤0.05;; *** p ≤0.001.

Article Snippet: The following antibodies were used: eIF2α (Abcam ab5369; 1:500), phosho-eIF2α[ser51] (Abcam ab32157; 1:500), GAPDH (Cell Signalling #2118; 1:5,000), β-actin (Cell Signalling #3700; 1:2500), eIF2Bα (Proteintech 18010-1-AP; 1:500), eIF2Bβ (Proteintech 11034-1-AP; 1:500), eIF2Bγ (Santa Cruz sc-137248; 1:500), eIF2Bδ (Proteintech 11332-1-AP; 1:50), eIF2Bε(Sigma-Aldrich HPA064370; 1:500).

Techniques: Western Blot, Expressing

Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: (A) Western Blot analysis of the level of eIF2Bα expression in U373, MO3.13 and SH-SY5Y cells following siRNA mediated silencing of eIF2Bα. Levels of eIF2Bα were normalized to levels of β-actin and presented as mean ± SD (n=3). p Values derived from an unpaired t test, ** p ≤ 0.01. (B) Average number of large eIF2Bα, eIF2Bβ, eIF2Bγ, eIF2Bδ, or eIF2Bε localized foci per cell following siRNA mediated silencing of eIF2Bα and/or 200 nM ISRIB treatment for 1h, presented as mean ± SD (n=3 counts of 30 cells) in (i) U373 cells, (ii) MO3.13 cells, and (iii) SH-SY5Y cells. p Values derived from a two-way ANOVA test, followed by a Tukey’s multiple comparisons analysis, * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, **** p ≤ 0.0001.

Article Snippet: The following antibodies were used: eIF2α (Abcam ab5369; 1:500), phosho-eIF2α[ser51] (Abcam ab32157; 1:500), GAPDH (Cell Signalling #2118; 1:5,000), β-actin (Cell Signalling #3700; 1:2500), eIF2Bα (Proteintech 18010-1-AP; 1:500), eIF2Bβ (Proteintech 11034-1-AP; 1:500), eIF2Bγ (Santa Cruz sc-137248; 1:500), eIF2Bδ (Proteintech 11332-1-AP; 1:50), eIF2Bε(Sigma-Aldrich HPA064370; 1:500).

Techniques: Western Blot, Expressing, Derivative Assay

Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: Western Blot analysis of the level of eIF2Bββ-ε expression in U373-MG, MO3.13 and SH-SY5Y cells following untreated and siRNA mediated silencing of eIF2Bα for 96h. Levels of eIF2Bββ-ε were normalized to levels of total protein stain and presented as mean ± SD (n=3). Data was analysed by two-way ANOVA test, followed by a Tukey’s multiple analysis. No significant differences observed.

Article Snippet: The following antibodies were used: eIF2α (Abcam ab5369; 1:500), phosho-eIF2α[ser51] (Abcam ab32157; 1:500), GAPDH (Cell Signalling #2118; 1:5,000), β-actin (Cell Signalling #3700; 1:2500), eIF2Bα (Proteintech 18010-1-AP; 1:500), eIF2Bβ (Proteintech 11034-1-AP; 1:500), eIF2Bγ (Santa Cruz sc-137248; 1:500), eIF2Bδ (Proteintech 11332-1-AP; 1:50), eIF2Bε(Sigma-Aldrich HPA064370; 1:500).

Techniques: Western Blot, Expressing, Staining

Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: (A) (i)Western Blot analysis of the level of eIF2α and eIF2α p[S51] expression and puromycin incorporation assays in U373, MO3.13 and SH-SY5Y cells following siRNA mediated silencing of eIF2Bα, 200 nM ISRIB treatment for 1h, or 1 µM Tg for 1h. (ii) Levels of phosphorylated eIF2α were normalized to levels of total eIF2α and presented as mean ± SD (n=3). (iii) Levels of puromycin were normalized to β-actin and are presented as mean ± SD ( n = 3). p Values derived from a two-way ANOVA test, followed by a Tukey’s multiple analysis, * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, **** p ≤ 0.0001. (B) ELISA analysis of the level of ATF4 expression in in U373, MO3.13 and SH-SY5Y cells following siRNA mediated silencing of eIF2Bα, 200 nM ISRIB treatment for 1h, or 300 nM Tg for 6h. Levels of ATF4 detected by ELISA are presented as mean ± SD (n=3). p Values derived from a one-way ANOVA test, followed by a Tukey’s multiple analysis, * p ≤ 0.05, ** p ≤ 0.01 *** p ≤ 0.001, **** p ≤ 0.0001. (C) Cells were transfected with Cy3 labelled siRNA negative control, Cy3 labelled siRNA targeting EIF2B1, and Cy3 labelled siRNA targeting EIF2B1 coupled with ISRIB 1h (200 nM) treatment. U373-MG and SH-SY5Y cells were fixed in methanol, MO3.13 cells were fixed in 4%PFA and subjected to ICC with anti-G3BP primary antibody and visualized using appropriate secondary antibodies conjugated to Alexa Fluor 488. Mean percentages of U373-MG, MO3.13 and SH-SY5Y cells with G3BPcontaining SGs. Error bars: ±s.d. Data was analysed using one-way ANOVA followed by a Tukey’s multiple analysis. *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001

Article Snippet: The following antibodies were used: eIF2α (Abcam ab5369; 1:500), phosho-eIF2α[ser51] (Abcam ab32157; 1:500), GAPDH (Cell Signalling #2118; 1:5,000), β-actin (Cell Signalling #3700; 1:2500), eIF2Bα (Proteintech 18010-1-AP; 1:500), eIF2Bβ (Proteintech 11034-1-AP; 1:500), eIF2Bγ (Santa Cruz sc-137248; 1:500), eIF2Bδ (Proteintech 11332-1-AP; 1:50), eIF2Bε(Sigma-Aldrich HPA064370; 1:500).

Techniques: Western Blot, Expressing, Derivative Assay, Enzyme-linked Immunosorbent Assay, Transfection, Negative Control

Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: A. Analysis of endogenous eIF2Bα and G3BP1 localizing to cytoplasmic foci in (i) U373; (ii) SKOV3 EIF2B1 WT/WT and SKOV3 EIF2B1 WT/L100P mutant cells following treatment with, 0.5mM H 2 O 2 or 500 µM RocA for 1h. Cells were fixed and subjected to ICC with anti-eIF2Bα (green) and anti-G3BP1 (red) primary antibodies and visualized using appropriate secondary antibodies conjugated to Alexa Fluor 488 and 594. Profile and surface plots of a representative focus show the level of colocalization (separate colours shown on graphics). Percentage of foci are presented as mean ± SD (n=3 counts in 30 cells with eIF2Bα localisation). Venn diagram of eIF2Bα and G3BP populations and co-localisation (n=3 counts in 30 cells with eIF2Bα localisation). B. Percentage of U373, SKOV3 EIF2B1 WT/WT and SKOV3 EIF2B1 WT/L100P cells with G3BP containing stress granules following 500 μM SA for 1h, 0.5mM H 2 O 2 or 500 µM RocA for 1h treatments (n=3 counts of 100 cells) . Mean percentage of cells displaying G3BP-containing SGs in a population of 100 cells per repeat. Error bars: ± s.d. (n=3). Data was analysed using one-way ANOVA followed by a Tukey’s multiple analysis. **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.000

Article Snippet: The following antibodies were used: eIF2α (Abcam ab5369; 1:500), phosho-eIF2α[ser51] (Abcam ab32157; 1:500), GAPDH (Cell Signalling #2118; 1:5,000), β-actin (Cell Signalling #3700; 1:2500), eIF2Bα (Proteintech 18010-1-AP; 1:500), eIF2Bβ (Proteintech 11034-1-AP; 1:500), eIF2Bγ (Santa Cruz sc-137248; 1:500), eIF2Bδ (Proteintech 11332-1-AP; 1:50), eIF2Bε(Sigma-Aldrich HPA064370; 1:500).

Techniques: Mutagenesis

Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: (A) Cells were treated with 125 μM SA for 30 minutes, 500 μM SA for 1h or 1 µM Tg for 1h and subjected to ICC with anti-eIF2Bα (green) and anti-G3BP1 (red) primary antibodies visualized using appropriate secondary antibodies conjugated to Alexa Fluor 488 and 594. Venn diagram shows total number of eIF2Bα and G3BP1 foci with the number of foci showing co-localisation (n=3 counts in 30 cells with eIF2Bα localisation). (B) Cells were treated with 500 μM SA for 1h and subjected to ICC with anti-eIF2Bε (green) and anti-G3BP1 (red) primary antibodies visualized using appropriate secondary antibodies conjugated to Alexa Fluor 488 and 594. Venn diagram shows total number of eIF2Bε and G3BP1 foci with the number of foci showing co-localisation (n=3 counts in 30 cells with eIF2Bε localisation). (C) Representative Airyscan images of (i) eIF2Bε and G3BP and (ii) eIF2Bα and G3BP following SA 1h (500 μM) treatments in SH-SY5Y and U373 cells respectively. eIF2Bε panel scale bar: 1 μM. eIF2Bα panel scale bar: 20 μM.

Article Snippet: The following antibodies were used: eIF2α (Abcam ab5369; 1:500), phosho-eIF2α[ser51] (Abcam ab32157; 1:500), GAPDH (Cell Signalling #2118; 1:5,000), β-actin (Cell Signalling #3700; 1:2500), eIF2Bα (Proteintech 18010-1-AP; 1:500), eIF2Bβ (Proteintech 11034-1-AP; 1:500), eIF2Bγ (Santa Cruz sc-137248; 1:500), eIF2Bδ (Proteintech 11332-1-AP; 1:50), eIF2Bε(Sigma-Aldrich HPA064370; 1:500).

Techniques:

Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: Confocal images of endogenous eIF2Bα and G3BP1 localizing to cytoplasmic foci in cells following treatment with 125 μM SA for 30 minutes, 500 μM SA for 1h or 1 µM Tg for 1h. Cells were subjected to ICC with anti-eIF2Bα (green) and anti-G3BP1 (red primary antibodies and visualized using appropriate secondary antibodies conjugated to Alexa Fluor 488 and 594. (A) U373 cells (B) MO3.13 cells (C) SH SY5Y cells

Article Snippet: The following antibodies were used: eIF2α (Abcam ab5369; 1:500), phosho-eIF2α[ser51] (Abcam ab32157; 1:500), GAPDH (Cell Signalling #2118; 1:5,000), β-actin (Cell Signalling #3700; 1:2500), eIF2Bα (Proteintech 18010-1-AP; 1:500), eIF2Bβ (Proteintech 11034-1-AP; 1:500), eIF2Bγ (Santa Cruz sc-137248; 1:500), eIF2Bδ (Proteintech 11332-1-AP; 1:50), eIF2Bε(Sigma-Aldrich HPA064370; 1:500).

Techniques:

Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: Analysis of endogenous eIF2Bα and G3BP1 localisation to cytoplasmic foci in (A) U373; (B) SKOV3 EIF2B1 WT/WT and (C) SKOV3 EIF2B1 WT/L100P cells following treatment with 500 μM SA for 1h. Cells were subjected to ICC with anti-eIF2Bα (green) and anti-G3BP1 (red) primary antibodies and visualized using appropriate secondary antibodies conjugated to Alexa Fluor 488 and 594. Profile and surface plots of a representative focus show the level of colocalization (separate colours shown on graphics). Percentage of foci are presented as mean ± SD (n=3 counts in 30 cells with eIF2Bα localisation). Venn diagram of eIF2Bα and G3BP populations and co-localisation (n=3 counts in 30 cells exhibiting eIF2Bα localisation). (D) Pymol diagram showing the interaction between eIF2Bα (red chain) and P-eIF2α (yellow chain). Missense mutations associated with neonatal diabetes (Franco et al., 2020) and the mutation present in the SKOV3 WT/L99P cells are highlighted

Article Snippet: The following antibodies were used: eIF2α (Abcam ab5369; 1:500), phosho-eIF2α[ser51] (Abcam ab32157; 1:500), GAPDH (Cell Signalling #2118; 1:5,000), β-actin (Cell Signalling #3700; 1:2500), eIF2Bα (Proteintech 18010-1-AP; 1:500), eIF2Bβ (Proteintech 11034-1-AP; 1:500), eIF2Bγ (Santa Cruz sc-137248; 1:500), eIF2Bδ (Proteintech 11332-1-AP; 1:50), eIF2Bε(Sigma-Aldrich HPA064370; 1:500).

Techniques: Mutagenesis

Journal: bioRxiv

Article Title: eIF2Bα subcellular localisation – a potential link between translation initiation and stress granule formation?

doi: 10.64898/2025.12.19.695478

Figure Lengend Snippet: Confocal images of endogenous eIF2Bα and G3BP1 localizing to cytoplasmic foci in U373, SKOV3 EIF2B1 WT/WT and SKOV3 EIF2B1WT/L100P cells following treatment with 500 μM SA for 1h, 0.5mM H 2 O 2 or 500 nM RocA for 1h. U373 cells were fixed in methanol and subjected to ICC with anti-eIF2Bα (green) and anti-G3BP1 (red primary antibodies and visualized using appropriate secondary antibodies conjugated to Alexa Fluor 488 and 594. The boxed region is enlarged, profile and surface plots were used to show colocalization (separate colours shown on graphics). Percentage of small and large eIF2Bα foci co-localising with G3BP were presented as mean ± SD (n=3 counts in 30 cells with eIF2Bα localisation). Venn diagram of eIF2Bα and G3BP populations and co-localisation (n=3 counts in 30 cells with eIF2Bα localisation).

Article Snippet: The following antibodies were used: eIF2α (Abcam ab5369; 1:500), phosho-eIF2α[ser51] (Abcam ab32157; 1:500), GAPDH (Cell Signalling #2118; 1:5,000), β-actin (Cell Signalling #3700; 1:2500), eIF2Bα (Proteintech 18010-1-AP; 1:500), eIF2Bβ (Proteintech 11034-1-AP; 1:500), eIF2Bγ (Santa Cruz sc-137248; 1:500), eIF2Bδ (Proteintech 11332-1-AP; 1:50), eIF2Bε(Sigma-Aldrich HPA064370; 1:500).

Techniques:

Journal: eBioMedicine

Article Title: A replicating RNA vaccine confers protection against Crimean-Congo hemorrhagic fever in cynomolgus macaques

doi: 10.1016/j.ebiom.2025.105698

Figure Lengend Snippet: repNP is immunogenic in cynomolgus macaques. Groups of cynomolgus macaques were sham-vaccinated or vaccinated with repNP-alone or repNP + repGc (a). CCHFV-specific IgG to whole-virus antigen (b) or recombinant antigen (c) was quantified by ELISA. CCHFV-specific IFNγ responses at day −14 to overlapping peptide pools spanning the CCHFV GPC (G1-14) or NP (N1-5) was measured by ELISpot (d). The isotype of CCHFV NP-specific antibody was measured (e) and the ability of NP-specific antibody to bind Fc-receptors, TRIM21 or complement component C1q measured (f). Line connects means (b,d,e,f). (c) Data presented as mean and SD.

Article Snippet: The bound antigen-specific antibodies were subsequently stained with PE-labelled tetramerized recombinant Fc receptors rhesus-FcγR2A (Duke Protein Production Facility, DPPF), rhesus FcγR3 (DPPF), human-FcγR2B (DPPF), human-FcRn (DPPF), human-C1q (Sigma C1740) and human-TRIM21 (Sino Biological 18010-H07B) for 1 h at room temperature, shaking at 800 rpm.

Techniques: Virus, Recombinant, Enzyme-linked Immunosorbent Assay, Enzyme-linked Immunospot

Journal: eBioMedicine

Article Title: A replicating RNA vaccine confers protection against Crimean-Congo hemorrhagic fever in cynomolgus macaques

doi: 10.1016/j.ebiom.2025.105698

Figure Lengend Snippet: Vaccinated animals develop anamnestic antibody responses to the NP and Gc. (a) IgG to recombinant NP, Gn and Gc was measured by ELISA at indicated timepoints relative to CCHFV challenge. (b) NP specific antibody was evaluated for its isotype and ability to bind Fc-receptors, C1q or TRIM21. Day 0 data is duplicated from Fig. 1 for comparison. (c) An IFNγ ELISpot was used to quantify anamnestic IFNγ responses to CCHFV peptides in PBMCs collected from animals at day 6 PI. (b,c) Line indicates mean.

Article Snippet: The bound antigen-specific antibodies were subsequently stained with PE-labelled tetramerized recombinant Fc receptors rhesus-FcγR2A (Duke Protein Production Facility, DPPF), rhesus FcγR3 (DPPF), human-FcγR2B (DPPF), human-FcRn (DPPF), human-C1q (Sigma C1740) and human-TRIM21 (Sino Biological 18010-H07B) for 1 h at room temperature, shaking at 800 rpm.

Techniques: Recombinant, Enzyme-linked Immunosorbent Assay, Comparison, Enzyme-linked Immunospot