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Prostate-specific antigen (PSA) is a glycoprotein enzyme and one of the most studied biomarkers for prostate cancer. The activity of PSA is well regulated and in the prostate, it is present as an inactive form. In a prostate tumor, enzymatically active PSA is present; and therefore, enzymatic activity can provide a quantifiable read out of malignancy in prostate cancer. PSA 750 FAST™ is a unique type of activatable fluorescent agent that can detect and quantify enzymatically active (PSA), and is also selective against unbound and complexed PSA.

PSA 750 FAST is optically silent upon injection and produces fluorescent signal after cleavage by enzymatically active PSA. Activation can only occur in the presence of active PSA. PSA 750 FAST may be used to monitor the progression of prostate tumors or to evaluate the potential therapeutic efficacy of drugs targeting the underlying mechanisms involved in these diseases.

Figure 1: Description and mechanism of PSA 750 FAST agent. In this case, the protease is PSA.

Figure 2: PSA 750 FAST (0.5 μM) was activated in the presence of various enzymes (0.1 μM activated enzyme) in the optimized buffer for each enzyme. Kinetic fluorescence readings were measured at 6 and 24 hours after adding enzymes. PSA 750 FAST is specifically cleaved by active PSA over inactive PSA (complexed with its inhibitor alpha1-antichymotrypsin) or other disease-relevant enzymes.

Figure 3: Pharmokinetics studies were performed using BALB/c female mice injected intravenously with 2 nmoles of PSA 750 FAST. Blood was drawn at the times shown on the graph. The plasma samples were analyzed by HPLC (UV 750 nm) and the half-life in normal mice was determined to be approximately 2 hours.

Products and catalog numbers

ProductCatalog NumberEx/Em wavelength (nm)Molecular weight (g/mol)Validated ExperimentsApplicationsStorage and Stability
PSA 750 FASTNEV11125687 (quenched), 750 (activated)/77042,500In vivo/Ex vivoOncology
Technical Data Sheet

Using PSA 750 FAST in vivo/ex vivo

  • The generally recommended procedure for in vivo imaging with PSA 750 FAST is administration via intravenous injection and imaging 6-24 hours post injection.
  • PSA 750 FAST will clear from tissues after approximately 50 hours. Repeat injection and imaging may be performed every 7 days for longitudinal studies. It is recommended that a pre-injection baseline image be taken prior to reinjection and imaging.
  • PSA 750 FAST enables imaging of PSA positive tumors such as LNCaP implanted in male nude mice.
ProductRoute of InjectionMouse Dose (25 g)Rat Dose (250 g)Blood t 1/2Tissue t 1/2Optimal imaging timeOptimal Re-injection Time (complete clearance)Route of Metabolism/ background tissueFMT and IVIS settings
PSA 750 FASTIV2 nmol6-20 nmol2 h60 h6-24 h
6-7 dBladderFMT 750/770
IVIS 745/800

In Vivo Imaging

Figure 4: Comparison of in vivo imaging by IVIS Spectrum and FMT2500. Imaging studies were performed in human prostate PSA+ LNCaP tumor-bearing male Nu/Nu injected intravenously with 2 nmoles of agent. The mice were imaged after 6 hours on the FMT2500 (in reflectance and tomographic modes) and on the IVIS Spectrum. PSA 750 FAST can be imaged efficiently and yields comparable results on both imaging systems.

Figure 5: Imaging studies were performed in human prostate PSA+ LNCaP tumor-bearing male Nu/Nu mice injected intravenously with 2 nmoles of PSA 750 FAST and imaged 24 hours later on the FMT2500. Eight different mice are shown in the above panel. PSA 750 FAST efficiently targets PSA+ tumors. Graphs show that the tissue half-life is approximately 2.5 days and that there is a good correlation between fluorescence signal from PSA 750 FAST and the tumor size.

Figure 6: Imaging studies were performed in tumor-bearing male Nu/Nu injected intravenously with 2 nmoles of PSA 750 FAST and imaged 24 hours later on the FMT2500. Four of the mice shown have LNCaP tumors (PSA+) and four have PC3 tumors (PSA-). Tumor fluorescence (in pmol) is shown in white next to the respective tumor (left panel). The panel on the right shows that the tumor fluorescence using PSA 750 FAST is significantly higher in PSA+ (LNCaP) than in PSA- (PC3) tumors in vivo.

Ex Vivo Applications

Figure 7: LNCaP (PSA+) tumor-bearing mice were injected intravenously with 10 nmoles of PSA 750 FAST. Twenty four hours later, tumors were excised and snap frozen in OCT media. Sections (ten micron) were prepared and the activated PSA 750 FAST fluorescence signal was visualized under fluorescence microscopy. PSA 750 FAST signal is shown in red and DAPI nuclear counterstain is shown in blue. PSA 750 FAST fluorescence is observed primarily in the periphery and center of the tumors.

Figure 8: LNCaP (PSA+) frozen tumor sections were incubated for 30 min with 5 mM PSA 750 FAST in the absence or presence of 5 mM of the natural inhibitor of PSA, alpha1-antichymotrypsin. PSA 750 FAST signal is shown in red and DAPI nuclear counterstain is shown in blue. PSA 750 FAST fluorescent signal significantly decreased by pre-incubating the tumor sections with the PSA natural inhibitor, a1-antichymotrypsin. This shows that PSA 750 FAST-associated signal is due to specific activation by enzymatically active PSA.

Frozen Tissue Protocol

We have validated PSA 750 FAST for use with frozen tissue samples (PSA+ tumors ex. LNCaP). Here is a brief protocol with a recommended concentration of agent to use:

  1. Freeze tumor (without agent) and section 5-10 µm by cryostat. For specificity, co-incubate sample with 5 µM alpha1-antichymotrypsin.
  2. Incubate with 5 µM PSA 750 FAST at 37 ºC for 30 min.
  3. Wash 1x with PBS.
  4. Mount with anti-fade reagent.
  5. Fluorescence microscopy filter: Cy5.5 or Cy7*


Please visit our Citations Library for references using PSA 750 FAST on the IVIS or on the FMT.